Urry D.W. (Ed.) What Sustains Life? : Consilient Mechanisms for Protein-Based Machines and Materials

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1.2 Four Principal Assertions of "What Sustains Life?''

1.2.3.3.2

Muscle Contraction as Inferred from

Model Protein Studies

• Stepwise, ATP binding to the cross-bridge

causes detachment from the thin filament by

bringing about dissociation of oil-like

domains.

• Binding of ATP also causes separation of

oil-like domains within the cross-bridge in a

way that allows forward extension of the

cross-bridge.

• Positive calcium ions pair with negative

vinegar-like groups associated with the thin

filament to uncover an oil-like domain for a

new cross-bridge attachment site.

• Phosphate release reestablishes oil-like

association between cross-bridge and thin fil-

ament and the power stroke occurs on re-

establishment of oil-like associations within

the cross-bridge.

This perspective, developed on applying the

elastic model protein studies to crystal struc-

tures of scallop muscle, brings us to the stage

longingly sought by Perutz in 1990 with the

statement "that one day we shall learn how

chemical energy is generated and turned into

motion."^^ Interestingly, at the time Perutz

penned these words, the demonstration of

chemical energy turned into motion by the

unexpected source of designed elastic-contrac-

tile model proteins of primary focus here had

been in the literature for little more than one

year.^^ Figure 1.7 gives the key results of that

publication in part A for acid-driven contrac-

tion at constant force (isotonic contraction) and

in part B for acid-driven force development at

constant length (isometric contraction).^^ The

fundamental process, regardless of the experi-

mental conditions, is acid-driven association of

oil-like domains.^^

The movable cusp of insolubility represented

in Figure 1.1 provides a means of visualizing the

molecular process. Addition of acid (proton,

H^) converts the vinegar-like carboxylate group

FIGURE 1.7. Shown are the first

reported data^^ of the conversion

by an elastic-contractile model

protein of chemical energy due

to an increase in concentra-

tion of acid into the mechanical

work of contraction. A Length

changes at constant force (iso-

tonic contraction) in phosphate-

buffered saline. B Force

changes at constant length

(isometric contraction) in

phosphate-buffered saline.

(Reproduced from Urry et al.^^)

•6.6 mm

time (hours)

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1 Iv

1 V L

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pH2.1 X^

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1 1 1

Si-

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20 40

time (hours)

60 80

Introduction

-COO"

to the more oil-like -COOH

group.

This

lowers the cusp of insolubility along the tem-

perature scale and results in association of oil-

like groups, which is a key element of the

contraction. When at fixed load, the association

(insolubilization) of oil-like domains lifts the

weight; when at fixed length, the association

(insolubilization) of oil-like domains stretches

interconnecting chain segments, resulting in an

increase in elastic force.^

1.2.3.4

Energy Conversion

in Mitochondria

The mitochondria are the energy factories that

produce ATP (adenosine triphosphate), the

energy currency of biology. In particular, a

single proton-driven protein-based machine in

the mitochondria produces 32 of the 36 ATPs

resulting from the oxidation to CO2 and H2O of

a single glucose molecule (the molecule that

best exemplifies a food source). This energy

conversion in mitochondria occurs in two steps:

(1) the oxidation of glucose-reduced carriers of

the electron transport chain that pumps

protons from one side to the other of the inner

mitochondrial membrane and (2) the flow of

protons back across the inner mitochondrial

membrane through a membrane-bound rotary

motor that drives the rotor extending into the

extramembrane rotary motor to produce the

ATP from ADP and

Pi,

inorganic phosphate.^^'^^

The doubled rotary motor is descriptively

named ATP synthase. The following four lists

of bullets provide succinct introductory state-

ments of this fundamental energy conversion of

biology.

1.2.3.4.1

The General Perspective

• ATP represents the energy coin of the bio-

logical realm.

• Mitochondria, biology's intracellular energy

factories, produce almost all of the ATP.

• Mitochondria produce ATP by a two-step

energy-converting process.

• The energies interconverted by both of these

steps are demonstrable by model proteins

designed to function by the consilient mech-

anism of controlling phase separation.

1.2.3.4.2

Step One for Energy Conversion in

Mitochondria as Interpreted from Model

Protein Studies

• In the first step at the cytosolic side of the

inner mitochondrial membrane, electron

removal forms a positively charged chemical

group with releasable protons.

• The positive charge disrupts hydrophobic as-

sociation, allowing proton release to that side.

• In a second step at the matrix side of

the inner mitochondrial membrane, electron

addition forms a negatively charged group

capable of proton uptake.

• The negatively charged group disrupts

hydrophobic association, allowing proton

uptake from the matrix side to result in net

transport of protons across the inner mito-

chondrial membrane.

1.2.3.4.3

Step Two for Energy Conversion in

Mitochondria in Two Parts

1.2.3.4.3.1 Part 1: The Fo-motor: Ten Hairpin

Loops of Protein Helices, with One Side of the

Loop Arranged as a Cleat of a Tractor

Tread,

Constitute a Transmembrane Rotary Motor

• Excess proton on one side of the membrane

enters a channel; in transit to the other side

the proton binds to vinegar-like group of a

transmembrane helix and makes it more

oil-like.

• The more oil-like helix, one of the ten protein

elements of a transmembrane rotary motor,

rotates into the oily layer of the membrane.

• The next helix rotates into position for a

half-

length channel site and releases its proton to

the opposite side of the membrane.

• This process rotates an oil-like double helical

rod that extends Uke an axle out of the center

of the wheel-like cluster of transmembrane

helices of the membrane-bound rotary motor.

1.2.3.4.3.2 Part 2: The Fi-motor: Transmem-

brane Rotary Motor Drives Rotation of Oil-like

Protein Rotor Within an Orange-Shaped, Six-

Subunit Protein Structure, Causing ADP and Pi

to Form ATP.

• The rotating axle extends to form the stem

and core of an orange-shaped protein struc-

1.2 Four Principal Assertions of "What Sustains Life?"

ture of six sections, with three symmetrically

arranged sections permanently bound to

ATP.

• The most oil-like side of the asymmetric

protein rotor, rotated by the Fo-motor,

begins to oppose one of three symmetrically

arranged catalytic sections that contains

ADP and Pi.

• Repulsion between the approaching most

oil-like side of the rotor and the most super

vinegar state of ADP and Pi produces ATP,

and complete exposure expels the ATP.

• Then ADP and Pi add to the emptied cat-

alytic site to form the most vinegar-like state

as that site faces the least oil-like side of the

rotor, and so on.

1.2.3.5

Molecular Aspects of the Fj-motor

The Fi-motor component of ATP synthase is

composed of a y-rotor and three a-subunits and

three p-subunits arranged as three aP pairs, that

is,

(aP)3.

Figure 1.8 schematically represents

three states of the Fi-motor in cross section.

Near their junctions with a p-subunit, each of

the three a-subunits contains one molecule of

ATP that is constant and does not change

during the catalytic process. Nonetheless, the a-

subunit ATPs fill an essential role in the con-

silient mechanism. From the crystal structure

data of Abrahams et al.,^^ the three P-subunits

are found with one catalytic site empty, a

second site contains ADP, and the third site

contains ATP. For purposes of our discussion,

ADP plus Pi (inorganic phosphate, which at

physiological pH would have as a dominant

species HP04"^) replaces ATP. The nucleotides

reside near the interfaces between the ap and

Pa pairs with the adenine component most

clearly defining its subunit, a or p. The polar

phosphate components reside at the interfaces

between the a- and P-subunits. In this case the

sides of the a- and P-subunits form the cleft that

emanates from phosphate to the y-rotor, as

sketched in Figure 1.8. Thus, the structural

feature of the charged phosphates of the

nucleotides residing at the base of a water-filled

cleft occurs with the Fi-motor just as it does

with the generalized ATPase considered above

and represented in Figure 1.6.

FIGURE 1.8. Illustration of

movable (rotating) cusp

of insolubility representing the cross section of the

Fi-motor of ATP synthase and depicting an asym-

metrical rotor with one of its three sides being very

oil-like. An oil-like association is shown to occurr

each time the most oil-like side of the rotor, rotated

by the Fi-motor, resides at an empty site. This pro-

vides a literal, spatial demonstration of the "movable

cusp of insolubility." On rotation of the oil-like side

of the rotor from the empty site to an ATP-filled or

an (ADP plus Pi)-filled site, the oil-like association

would be lost. The metaphorical representation

becomes the "movable cusp of insolubility" with

the graphical equivalence of the leftmost (low-

temperature) cusp of Figure 1.1 being raised to

the rightmost (high-temperature) cusp. Thus, the

"movable cusp of insolubility" at which biology

thrives can, by the Fi-motor of ATP synthase, be

given both literal and metaphorical representation.

Introduction

1.2.3.6

The Fi-Motofs Cusp

of Insolubility in Both Assembly

and Function

1.2.3.6.1

When Assembled, the Fi-motor

Functions as a Rotating Cusp of InsolubiUty

The "movable cusp of insolubility" of the Fi-

motor becomes both literal and metaphorical.

The metaphorical representation occurs as the

oil-like side of the rotor passes from association

with the oil-like empty P-subunit where the

cusp of insolubility, represented in Figure 1.1,

resides at low temperature (below physiologi-

cal temperature) to high temperature, that is,

above physiological temperature to result in

oil-like dissociation of rotor from sleeve. Thus,

the metaphorical representation occurs as the

oil-like side of the y-rotor is rotated by the

FQ-

motor from association with an oil-like empty

P-subunit to dissociation from a vinegar-like a-

subunit that contains ATP.

For the appropriate concentrations of ADP

and Pi, Figure 1.8 demonstrates a Hteral visual

representation of going from association to

dissociation and back to association for each

120° rotation. This literal "movable cusp of

insolubility" results from the competition for

hydration that exists between oil-like and

vinegar-like groups as in "oil and vinegar don't

mix.'' Expressed as a repulsion, the same com-

petition for hydration occurs involving interac-

tion of the super vinegai-hke group of ATP and

the most super vinegar-like state of ADP plus

Pi with the oil-like side of the y-rotor in Figure

1.8. An analogous situation occurred between

the very oil-hke phenylalanine groups of

Model protein v in Figure 1.4 and the classic

vinegar-hke carboxylate group with its single

negative charge. In the latter case the repul-

sion was 2.4kcal/mol. In Figure 1.8, the repul-

sion would be many times greater as the

nucleotide phosphates would have more nega-

tive charges and the change in the oil-like char-

acter of the three faces that define the central

y-rotor are so striking, namely, -20, 0, and +9

kcal/mol (see Chapter 8, section 8.4.4.3, and

Table 8.2).

1.2.3.6.2

The Cusp of Insolubility for the

Fi-motor, Ahas the Fi-ATPase When Running

in Reverse

When the rotor of ATP synthase is driven

clockwise by the Fo-motor, as shown in the pro-

gression from states A to B to C in Figure 1.8,

synthesis of nearly 90% of biology's ATP

occurs. When the Fj-motor is separated from

the Fo-motor, it can function in reverse as an

"idling" rotary ATPase called the Fi-ATPase,

or it can have a filament attached to it and

perform the work of rotating the filament.^^ As

occurs with many enzymes, Fi-ATPase cold

denatures,^^ that

is,

its subunits—the y-rotor and

an accompanying e-subunit together with the

three aP components within which the rotor

rotates—dissociate on lowering the tempera-

ture.

In other words, the oil-hke associations

between the subunits of Fi-ATPase separate on

lowering the temperature and recombine on

heating to physiological temperature. There-

fore,

the left-hand cusp of Figure 1.1 represents

the cusp of insolubility for the oil-like assem-

bly/disassembly of the functional Fi-motor.

1.2.3.7

Considered Steps in the Fj-motor

Synthesis of ATP as Shown by the

Consilient Mechanism

1.2.3.7.1

Consilient Steps in the Synthesis of

ATP by the Fi-motor

The Fi-motor structure due to the Walker

group^^ functions ideally to demonstrate the

steps whereby the rotating cusp of insolubility

would result in the synthesis of

ATP.

The y-rotor

may be represented by the relative oil-like

character of its three faces that are calculated

in Chapter 8, section 8.4.4.3, shown in Figures

8.31 to 8.33, and listed in Table 8.2. As repre-

sented in Figure 1.8, there is a very oil-like face

(marked by -20 and shown directed toward the

empty catalytic site), a neutral face (indicated

by 0 and directed toward the p-ADP-hPi site),

and a relatively vinegar-like face (indicated by

+9 and directed toward the site indicated in

Figure 1.8 as occupied by P-ADP).

1.2 Four Principal Assertions of "What Sustains Life?"

Starting with the structure and occupancy

states of Figure 1.8 and given the insight of the

consiUent mechanism, Pi would add to the

p-ADP site while facing the +9 side of

the y-rotor and would do so as the -20 face of

the y-rotor began clockwise rotation. Then the

y-rotor, driven by the Fo-motor, would continue

rotation in a clockwise direction until the very

oil-like, -20 face would begin opposing the P-

(ADP plus Pi)

site.

As the repulsion between the

oil-like side of the rotor and the most super

vinegar-like (ADP plus Pi) state would build,

the repulsion would be relieved by formation of

the lesser but yet super vinegar-like ATP

product. On continuing the 120° rotation to

complete the face-off against the most oil-like

side of the rotor, this most oil-like face of the y-

rotor would force the ATP product out of the

catalytic site. As the oil-Uke face of the rotor

begins rotation from the recently emptied site,

ADP and Pi could enter with the least oil-like

face

(-1-9)

in apposition, and the beginning of the

next 120° rotation would bring in the interme-

diate oil-like face (0) to close the site before

facing off with the most oil-like face (-20).

1.2.3.7.2

The Functional Role of the Three

Symmetrically Arranged, Noncatalytic,

ATP-containing a-subunits

The ATP-containing a-subunits are arranged

with threefold symmetry around the y-rotor. By

the consilient mechanism, the a-ATP molecules

become a trigonal force array focused on the

central y-rotor that would serve to keep the

asymmetrical rotor more central and would

serve to prevent insolubilization involving a

more oil-like side of the rotor and the sleeve.

As required for efficient function, the three a-

ATP molecules would assist in preventing a

viscous drag or a freezing-up by hydrophobic

association between rotor and the central parts

of the a- and p-subunits. Accordingly, by the

consilient mechanism of water-mediated repul-

sion between oil-like elements of the y-rotor

and the super vinegar-like phosphates of the a-

ATP molecules, the ATP-containing noncat-

alytic sites form a crucial function by providing

a structure capable of a more efficient energy

conversion.

1.2.3.8

The ''Waters of Thales''Within the

Fj-motor of ATP Synthase

To be workable, the consihent mechanism of

repulsion between oil-Uke and vinegar-like

(e.g., charged) groups, technically called an

apolar (oil-like)-polar (e.g., charged) repulsive

free energy of hydration, requires the presence

of adequate water to form between oil-like and

vinegar-like groups. Therefore, the first test

of relevance of the consilient mechanism is

to demonstrate sufficient water within the

protein-based machine. As considered above

for the myosin motor. X-ray diffraction

methods for protein structure determination

have reached the stage where at least some of

the water molecules in the structure can be

seen. As the observed water molecules must be

fixed in position during the long period of time

required for collecting the X-ray diffraction

pattern, more mobile water molecules are not

seen. Thus any observed water molecules rep-

resent a minimal number of the actual water

molecules present.

A top view of the Fi-motor is shown in cross-

eye stereo view in Figure 1.9A, where the three

aP pairs are given in space-filhng representa-

tion and the y-rotor and its attached e-subunit

are given in a backbone-only representation.

The light dots are the water molecules observ-

able on the surface of the Fi-motor. When seen

in stereo, the observed water molecules almost

entirely reside in clefts and recesses of the

surface that in reality are totally filled with

water molecules. This provides an opportunity

to obtain a sense of how few water molecules

are observed out of those actually present.

Showing exactly the same view, but with the

three pairs of ap-subunits removed, the stereo

view in Figure 1.9B, allows the number of

observed water molecules within the Fi-motor

to be seen. Tlierefore, these observed water

molecules become the "waters of Thales"^^'^"^

required to give this most important protein-

based machine of biology the capacity to func-

tion.^"^ Living organisms are "kept alive" by the

"waters of Thales"^^; they are the waters

through which the repulsive force between

oil-like and vinegar-like groups becomes

expressed.

Introduction

i4«

FIGURE 1.9. Observation of the "waters of

Thales"

the Fi-motor of ATP synthase. The crystal structure

of the Fi-motor of ATP synthase is shown in cross-

eye stereo view from the top axis having all sites

filled with ATP and with the rotor given by the fine

lines of a backbone representation. A A space-

filling model of (ap)3 shows detected surface waters.

B A space-filling view of all water molecules

detected by X-ray diffraction demonstrates the inter-

nal "waters of Thales" available to function by the

consilient mechanism for protein-based machines.

(Figure preparation was based on the crystallo-

graphic data of Abrahams et al.^^ obtained from the

Protein Data Bank, Structure File IBMF.)

Immediately above, v^e have noted key

stages of the consilient basis for the energizing

and de-energizing processes. In Chapter 2, v^ith

all sites containing nucleotide, a slightly more

detailed illustration of the ATP-forming part of

the rotary motor than in Figure 1.8 adds further

structural perspective of a cusp of insolubility

that is externally driven to rotate. In Chapter 8,

and in a separate publication,^^ arguments at

molecular detail address this remarkable rotary

motor that synthesizes most of the energy cur-

rency required by living systems.

1.2

Four Principal Assertions

"What Sustains Life?'"

1.2.3.9

Hemoglobin Transport

Oxygen from

the

Lungs

to the

Tissues,

Two

Parts

Immediately below,

note

the

historical case

of oxygen transport

hemoglobin from lungs

the

tissues

and

note

how

this would

dis-

cussed

terms

of the

consilient mechanism.

This

was the

first biological system appreciated

for

its

positive cooperativity,

the

cooperative

binding

oxygen molecules that

is key to

suc-

cessful transport.

1.2.3.9.1

Part

Happenings

in the

Arterioles

the

Lungs

•

In the

lungs,

the

first

four vinegar-like

ox-

ygens binds poorly

to one of

four nearly iden-

tical units comprising hemoglobin because

oxygen binding disrupts transient hydration

that maintains oil-like domain association.

•

The

second oxygen binds more easily

there

is now

less oil-Hke hydration

disrupt.

•

The

third

and

fourth oxygens bind more easily

yet,

complete loading

of the

hemoglobin

transport system with

its

oxygen cargo.

• Thus nearly empty hemoglobin entered

the

lungs where there

enough oxygen

load

fully

the

hemoglobin molecule, which leaves

the lungs

transit

to the

tissues.

1.2.3.9.2

Part

Happenings

in the

Arteriovenous Junction

of the

Tissues

•

In the

tissues,

CO2 and

biphosphoglycerate,

a super vinegar-like molecule with many neg-

ative charges, bind

positive sites,

the

latter

with many positive charges where four

sub-

units

hemoglobin meet.

• Association

many negative with positive

charges neutralizes their charge effect

and

results

in a

shift toward more

oil

hydration

and

its

consequence

dominance

oil-like

domains.

•

The

increase

in oil

hydration shifts toward

release

vinegar-like oxygen,

and

spatially

localized oil-like domains between protein

subunits associate.

•

this way, hemoglobin provides

efficient

oxygen transport vehicle

being able

release more

of its

load

in the

tissues.

1.2.3.10

Hemoglobin Mutation

of a

Single

Amino Acid from Vinegar-like

Oil-like

Causes Dysfunction

Shifting

the

Cusp

of Insolubility

Too Far

Toward Insolubility

•

The

disease sickle cell anemia provides

another example

coherent phenomena.

• Sickle cell anemia

is due to a

mutation

of a

vinegar-like

to an

oil-like group

on the

surface

hemoglobin.

•

The

resulting increase

in oil

hydration poises

the hemoglobin molecules

for

aggregation.

• Limited oxygen supply allows

shift toward

sufficient

oil

hydration

trigger hemoglobin

phase separation

and the

onset

of the

symp-

toms

of the

disease.

1.2.3.11

Coherence Between Assertions

and 3 and

Monod's ''Second Secret

of Life"

As noted

Perutz,^'^^ Monod beUeved that

his

discovery

allosteric effects,

the

existence

two

more states

equilibrium that give rise

the

efficient function

positive cooperativ-

ity, constituted

the

"second secret

life,"

second only

to "the

structure

of DNA."

Monod's discovery

was

exciting;

represented

a phenomenological observation,

axiom,

describable

by a

mathematical formalism.

The phenomenological observation

and the

associated mathematical representation were,

however, devoid

of a

physical basis

for the

con-

version between structural states.

The

Mecha-

nistic Assertion

this book fills that void.

Furthermore, Monod impugned

the

simple

problem

translational symmetry

fibrous

structures

to be "set

aside"

as not

having rele-

vance

to "the

second secret

life."

Thus,

find

the lauded allostery

model proteins designed

from

repeating sequence

of the

fibrous

protein elastin

was

unanticipated

in the

com-

munity

large. Furthermore, these elastic-

contractile model proteins have been designed

to perform

the

family

energy conversions

extant

biology. Therefore,

the

remarkable

finding of coherence

phenomena

positive

cooperativity between designed elastic-

contractile model proteins

and

biology's

proteins requires that

the

mechanism whereby

Introduction

these model proteins perform their diverse

functions be given serious consideration for

function of biology's proteins.

The Hill plots in Figure 1.3B compare the

Hill coefficients of hemoglobin and myoglobin.

The celebrated positive cooperativity of hemo-

globin exhibits a Hill coefficient, n, of 2.8. This

positive cooperativity evolved for the efficient

transport of oxygen from lungs to the tissues.

On the other hand, myoglobin, with the role of

storage and diffusion of an innocuous state of

oxygen in the tissues, exhibits a Hill coefficient

of 1, indicating the presence of neither positive

nor negative cooperativity. As will be shown in

Chapter 7, the replacement in myoglobin of

many of protein's most vinegar-like residues,

the carboxylate-containing glutamic acid (Glu,

E) residues, by more oil-like residues results in

the oil-like association of four myoglobin-hke

chains, two a-chains and two |3-chains, to form

hemoglobin, designated as a2p2- The allosteric

structural change on oxygenation occurs at the

oil-like interface between pairs of ap dimers. In

our view, this change in oil-like association at

the a^P^-a^p^ interface is responsible for the

positive cooperativity of hemoglobin.

Larger Hill coefficients indicate more effi-

cient function. Interestingly, the set of designed

elastic-contractile model proteins in Figure

1.3A exhibit Hill coefficients with an extraordi-

nary range from 1.5 to 8.0. Thus, these de

novo-designed elastic-contractile model pro-

teins exhibit a comprehensible range of effi-

ciencies that depend on the degree of oil-like

character of the model protein and that result

from the competition for hydration between

oil-like and vinegar-like functional groups, as

developed in Chapter 5. It is not inconsequen-

tial,

therefore, that the understanding devel-

oped as the mechanistic assertion of this book

gives rise to parameters for measuring effi-

ciency that can exceed those widely acclaimed

in the function of biology's multisubunit

proteins.

Thereby, the objective of this book is to

demonstrate the phase separation mechanism of

oil-like domains separating from water and

related controlling phenomena at the molecular

level as relevant to specific biological protein-

based machines.

12A Assertion

The Applications

Assertion

1,2,4,1

The Comprehensive Capacity to

Control Association of Oil-like Domains

Provides the Necessary Understanding for

Meaningful Engineering of Protein-based

Materials for Diverse Medical and

Nonmedical Applications

The families of model proteins by which the

fundamental mechanism is illustrated become

designable materials with potential for sustain-

ing both individual health and an increasingly

complex and populous society. This includes all

polymers operating under the consilient mech-

anism where adequate control over composi-

tion and sequence exists.

The first assertion, as presented in Chapter 5,

results in five rules (axioms) for the design of

model proteins for energy conversion by the

phase separation mechanism. We began these

developments with a simple hypothesis visual-

ized as cusps of insolubility, as shown in Figure

1.1: Any change that makes the elastic model

protein more oil-hke lowers the onset tempera-

ture for association of oil-hke domains, that is,

lowers the temperature for the cusp of insolu-

bility. Similarly, any change that makes the

elastic model protein more vinegar-like raises

the onset temperature for association of oil-like

domains, that is, increases the temperature

range for the cusp of insolubility. On this basis

we systematically designed model proteins with

varying composition for the purpose of achiev-

ing different energy conversions. Testing of

these designed model proteins indeed demon-

strated the intended energy conversions. Five

phenomenological axioms for energy conver-

sion resulted. In the fourth assertion these rules,

buttressed by development of an understanding

of the underlying mechanism (the second asser-

tion) and by the demonstration that biology's

protein-based machines function using this

mechanism (the third assertion), substantiate

the blueprint for designing new model proteins

to fill needs in health care and to support an

increasingly complex and populous society.

In the design focus of the conventional archi-

tect, gravity is the force of primary concern. For

1.2

Four Principal Assertions

"What Sustains Life?'

architectural design

at the

molecular level

the aqueous milieu

biology, however,

the

forces associated with

the

separation

of oil

from

vinegar

and of oil

from water dominate.

The

medium

for

design

at the

molecular level does

not involve

the

steel, cement, brick, mortar,

wood, glass,

even petroleum-based plastics

the traditional architect,

and it is not the

closely related petroleum-based material

of the

typical bioengineer

and

biomaterials scientist.

Rather,

our

medium

for

design

is the

same

that

biology

itself,

protein-based materials

water. Skillful

use of the

forces controlHng

protein folding, assembly,

and

function allows

for

the

design

medical

and

other useful

devices

and

materials.

The approach

of the

fourth assertion

is one

of utilizing biology's

own

natural processes

solve health-care

and

broader societal prob-

lems.

This inherent need rings

in the

concluding

paragraph

of E.O.

Wilson's book Consilience:

The Unity

Knowledge, where

states

"To

the extent that

depend

prosthetic devices

to keep ourselves

and the

biosphere ahve,

will render everything fragile. To

the

extent that

we banish

the

rest

life,

will impoverish

our

own

species

for all

time."^^

There

are as yet no

commercial sales

elastic

and

plastic protein-based polymers,

but

recent progress

correcting relevant intellec-

tual property rights

by the U. S.

Patent

and

Trademark Office will hopefully allow

for

com-

mercial uses

in the

near future. Thus,

the

appli-

cations noted below must

yet be

seen

terms

potential rather than

realized utility.

1.2.4,2

The

Remarkable Biocompatibility

of These Elastic Model Proteins

• These elastic model proteins exhibit remark-

able biocompatibihty

due to an

elasticity

dif-

ferent from that

disordered petroleum-

based elastomers

and

natural latex rubbers.

• These elastic model proteins form regular

dynamic structures that exhibit mechanical

resonances

at low

frequencies.

•

approach

and

identify these elastic mate-

rials

foreign requires direct interaction,

for

example, with antibody.

•

The

mechanical resonances present

barrier

to interaction,

their coherent motions must

be stopped

order

for

antibodies

interact

with

the

biomaterial

and to

identify

it as

foreign.

1.2.4.3

The

Design

Medical Care

Devices

for

Soft Tissue Restoration

1.2.4.3.1

Prevention

Adhesions

and

Correction

Urinary Incontinence

•

The

soft tissue

mammals, from which

our

model proteins were developed,

is a

compU-

ant tissue.

• Consequently, soft tissue restoration

be-

comes

manifest area

application

elastic model proteins.

• Preventing scar tissue formation after spinal

surgery holds promise

correct

the

problem

of failed back surgery syndrome

and its

con-

sequence

painful disability.

• Inducing tissue generation

at the

base

of the

bladder

can

restore urinary continence from

a state that constrains activity

and

limits

quality

life.

1.2.4.3.2

Function/Adaptive-Restructuring

Cycles

Tissue Restoration

•

At the

cellular level, function/adaptive-

restructuring cycles

can be

quite explicit.

• Cells

in an

arterial wall have multiple attach-

ments

surrounding connective tissue.

• Through these attachments, arterial cells

become stretched

and

relaxed

blood

pulses through

the

artery.

• Stretching/relaxing provides

the

energy stim-

ulus that transforms into

chemical energy

output

for

remodeling

arterial wall

order

better sustain

the

pulsing forces.

1.2.4.3.3

Restoration

Arteries

• Tissue restoration

can

utilize this principle

function/adaptive-restructuring cycles.

• Synthetic elastic tubes formed from elastic

model protein

can

match

the

elastic proper-

ties

of the

natural artery.

•

The

synthetic elastic tubes

can

also contain

cell chemoattractant

and

attachment sequ-

Introduction

ences

provide attachment sites

for the

natural arterial cells.

•

such temporary functional scaffoldings,

natural cells

can be

induced into

the

scaffold,

attached,

and

stretched

due to the

pulsing

blood;

as a

consequence, cells remodel

the

synthetic scaffold into

natural tissue suffi-

cient to sustain essential mechanical function.

1.2,4.4

The

Design

Medical Care

Devices

for

Controlled Drug Release

1.2.4.4.1

Drug Delivery

by the

Phase

Separation Mechanism

• From loading

release, drug delivery

can

be achieved

by the

phase separation

mechanism.

• Drugs, containing charge, pair with oppo-

sitely charged vinegar-like groups

of the

elastic model protein.

• Increase

in oil

hydration

due to the

asso-

ciation

opposite charges causes phase

separation.

• The separated phase

designed protein plus

drug constitutes

the

drug delivery vehicle.

1.2.4.4.2

Increasing Oil-like Character

Charged Model Protein Enhances

Ion

Pairing

• Above

protein design,

the

drug provides

the chemical energy

for its own

packaging.

•

The

protein

can be

designed with

range

oil-like groups while maintaining charge.

•

The

more oil-like association

pairing

oppositely charged drug

and

model protein,

the more tightly

the

drug

held.

•

The

more tightly held

the

drug,

the

slower

the release.

1.2.4.4.3

Strength

of Ion

Pairing Controls

Rate

Drug Release

•

large number

drugs have

charged,

vmegar-like part

and an

oil-like part.

• Examples

are the

many analgesics

and

anes-

thetics (painkillers), including narcotics.

•

The

loaded drug delivery vehicle

can be

implanted

in the

body.

•

design

model protein, release from

the

implant

can be

sustained

at a

desired level.

1.2.4.4.4

Controlled Release

Narcotic

Antagonists from Subcutaneous Implants

• There

are

also narcotic antagonists that block

the action

drugs like heroin

and

cocaine.

•

The

drug delivery vehicles containing nar-

cotic antagonists

can be

implanted

in the

body.

• Release from

the

implant would block,

for

example,

the

action

of a

large dose

heroin.

•

The

potential

is for

sustained dehvery

narcotic antagonist,

high,

redepen-

dence,

and the

delivery vehicle

can be

designed

gracefully disappear

as its

task

completed.

1.2.4.5

The

Design

Medical Care

Devices

for

Vaccine Delivery

Nanoparticles, with different affinities

for

vaccine

and

with potential

to be

adjuvants,

means

of a

single inoculation could replace

spaced multiple immunizations.

•

The

drug delivery vehicle

can be

formed

charged nanoparticles

different oil-like

composition with

the

potential

for

function-

ing

as an

adjuvant.

•

The

different nanoparticles would take

vaccine with different binding affinities

and

different release profiles.

•

The

vaccine-loaded nanoparticles could

introduced into

the

body through epithelial

lining without

the

need

for

injection.

•

The

requirement

for

multiple vaccinations,

spaced over many months, could

replaced

with

single treatment using

family

nanoparticles each with

different release

level

and

period

function

multiple

immunizations.

1.2.4.6

Design

Nonmedical

Applications: Release of Agricultural

Enhancement Factors

Controlled release

can

enhance factor effec-

tiveness

and

limit harmful environmental side

effects.

• Controlled release

agricultural enhance-

ment factors

analogous

drug delivery.