Download Some General Physical and Chemical Properties of Proteins

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SOHE GENERAL PHYSICAL A N D CHENICAL
PROPERTIES OF PROTEINS
The large c l a s s of compounds now c l a s s i f i e d as "pmteins" represent a polycomplexity of individual species, yet, a l l have many common
c h a r a c t e r i s t i c s , are constructed f r o m similar chemical u n i t s and exhibit
similar physical and chemical properties. Proteins are biosynthesized f r o m
a v a r i e t y of chemical molecules which a r e broadly c l a s s i f i e d as a-amino
acids. As the synthesis proceeds and the protein chain i s formed, the
a-amino acids a r e caref'ully selected and one added t o another i s a s p e c i f i c
manner u n t i l the growth process i s terminated. Nature i s s p e c i f i c i n the
s e l e c t i o n of the correct amino acid at the t i m e it i s needed i n the format i o n of t h e polymer chain.
The chemical c h a r a c t e r i s t i c s of the proteins depend upon the individual amino acids u n i t s present i n the complete protein molecule and t h e
sequence i n the u n i t s as they are placed i n the polymer chain. That is, t h e
chemical properties w i l l not only be influenced by which amino acid has ent e r e d t h e chain but a l s o upon i t s nearest neighbors. However, some of t h e
physical properties w i l l be r e l a t i v e l y independent of the chemical constmct i o n and will r e s u l t mostly from the fact that proteins j u s t as o t h e r synt h e t i c high molecular weight compounds, are i n essence, giant molecules
whose physical behavior i s memly a r e f l e c t i o n of t h e i r enormous s i z e .
The chemical u n i t s of constmction, the a-amino acids, have many
distinguishing f e a t u r e s and are schematically represented by t h e following
formula;
H'
N
'H
OH
vhere the starred carbon i s asymmetric and thus exists i n two, o p t i c a l
isomeric forms, the right (d) and l e f t (1)hand molecules. Since nature
produces and u t i l i z e s only a-srrcino acids i n t h e l-form the joining of d and
1 u n i t s does not become a complication i n protein structure.
The R group attached t o the asymmetric carbon atom i s responsible
f o r the d i s t i n c t i o n of one pamino acid from another. R can be either pure
hydrocarbon i n chemical nature o r may contain o t h e r elements such as
oxygen and nitrogen carboxylic a c i d o r amino groups.
However, the simple molecule described above has other more int e r e s t i n g features since it contains both a basic substituent (-NHz) and an
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a c i d i c substituent (-CCOH) making the amino acids rather unique molecules
In basic solution the carboxylic acid portion would be more completely
Ionized wheE the molecule would take a structure indicated by 11, thus
becoming an anion, and
H
0
in a c i d i c solution a s t r u c t u r e as shown i n I11 r e s u l t s where the amino a c i d
be come 8
0
Consequently i n an e l e c t r o l y t i c c e l l the migration of an amino
a c i d t o the negative electrode (cathode) o r the positive electrode (anode)
w i l l depend not dnly on the individual amino a c i d but the pH of the media i n
which it I s dissolved.
a cation.
The ability of 8 molecule t o behave both as a base and a6 an a c i d
is termed amphoterism and the molecules ampholytes. This phenomena i s not
limited t o amino-acids but is also of property of many simple metal hydroxides.
The acid strength of an amino a c i d i n aqueous solution w i l l depend
p r i n c i p a l l y on t h e substituent R. Depending on the s t r u c t u r e and composition
of R s o w amino acids i n aqueous solution w i l l be acidic in nature (pH < 7 ) ,
Amino acids of this type a r e weak acids and a portion of the carboxylic acid
groups i n t e r a c t wIth water and dissociate as indicated below (IV).
Other amino acids w i l l i n t e r a c t with water at t h e amine s i t e a8 shown in (V),
H
1
I
,
0
H
I
0
and thus give basic aqueous solutions. S t i l l others react equivalently with
water as i n IV and V SO that no net change i n s t r u c t u r e r e s u l t s and the amino
acid i s e s s e n t i a l l y neutral i n nature.
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By proper adjustment of the pH of an aqueous solution of amino
acids LL point is reached where the net e l e c t r i c a l charge on t h e amino acid
is zero. Under these specific conditions the molecule would not migrate t o
either electrode under the influence of an e l e c t r i c field. The pH where
t h i s phenomena is exhibited is termed the i s o e l e c t r i c point and is a chara c t e r i s t i c physical property of a l l amino acids. IWy o t h e r physical prope r t i e s such as membrane potential, o p t i c a l rotation, s o l u b i l i t y , diffusion,
s t a b i l i t y and resistance t o denaturation w i l l show a maxima o r minima a t
t h i s unique pH.
When proteins a= formed the amino acids a c t as difunctional
monomers and polycondense by eliminating water t o form amide linkages and a
polypeptide chain of the nature i l l u s t r a t e d by(V1)
Although the amine and carboxylic a c i d groups are destroyed i n t h e peptide
fomation, t h e protein s o formed may r e t a i n free amino o r acid groups as
dangling side groups (pendant groups as embodied in R) o r as terminal unreacted end groups. However, even though the peptide linkage will no longer
take part i n pH t i t r a t i o n s a l l residual amine and acids groups w i l l . Thus
proteins are Ellso mpholytes and exhibit an i s o e l e c t r i c point where properties such as Solubility, solvent swelling snd molecular size i n solution
w i l l be d r a s t i c a l l y affected by slight a l t e r a t i o n of the pH. In f a c t , t h e
i s o e l e c t r i c point can be more sharply defined f o r proteins than f o r many of
i t s contributing a-amino acids when the f o m r is measured by t h e T i s e l i u s
electrophoresis method.
In t h i s method a dissolved aqueous solution of protein is placed
i n an e l e c t r i c f i e l d and the pH of the solution is changed by t i t r a t i o n
with an acid o r base. The moving boundary of ions can be followed by s u i t able instrumental methods and when the boundary ceases t o move in the f i e l d
the molecule i s i n the i s o e l e c t r i c s t a t e .
Many of t h e physical c h a r a c t e r i s t i c s of proteins depend upon
t h e i r s t r u c t u E and the f a c t t h a t they a m giant molecules. Structure wise,
the proteins as a class generally have regions of high c r y s t a l l i n i t y . However, as i n a l l long chain molecules it is impossible f o r proteins of high
molecular weight t o completely c r y s t a l l i z e . This r e s u l t s f r o m the complications involved i n making a molecule composed of many segments rearrange
each segment i n a specific order within the chain and with respect t o its
nearest neighbor molecules i n a reasonable period of time. Thus proteins
which c r y s t a l l i z e will contain both crystalline regions where high l o c a l
order persists, and amorphous regions where very l i t t l e order e x i s t s . It
is these l a t t e r regions which are the most vulnerable t o the action of
solvents.
These types of protein molecules behave somewhat as synthetic network molecules such as Vulcanized rubber and w i l l reach an equilibrium
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degree of swelling providing the imbibed solvent-polymer interactions a r e
t o o weak t o overcome the forces of c r y s t a l l i n i t y . The c r y s t a l l i n e content
of proteins can be both enhanced and destroyed by s u i t a b l e changes i n pH
o r metal ion strength of imbibed water. Some proteins whose c r y s t a l l i n i t y
has been destroyed by dissolution w i l l f a i l t o r e c r y s t a l l i z e a f t e r removel
f r o m t h e solvent and are thus considered denatured o r a l t e r e d i n an irrev e r s i b l e fashion.
While some protein molecules are joined together by a mutual
sharing of some of their segments i n a c r y s t a l l i t e , others are cross
linked i n t e m i t t e n t l y by covalent chemical bonds. This type of r e s t r i c t i o n tends t o reduce the symmetry of a protein chain thus r e s u l t i n g i n low
degrees of c r y s t a l l i n i t y . Often these cross l i n k s can be s e l e c t i v e l y
broken and the r e s u l t i n g chains solubilized. However, as cross l i n k s they
are network junction points and g r e a t l y influence c e r t a i n physical propert i e s of the proteins (such as equilibrium swelling, modulue of e l a s t i c i t y ,
etc.).
One of the properties affected i s the degree t o which a network
may be swollen by imbibing solvent. With t h e cross l i n k s a c t i n g as
r e s t r i c t i o n s the polymer w i l l swell with solvent u n t i l the elastic retract i v e energy of the chain reaches a state of equilibrium with energy res u l t i n g from solvent-protein interactions. The equilibrium degree of
swelling should be reproducible once the conditions f o r equilibrium are
reached and providing no degradation of protein takes place.
Several f a c t o r s a f f e c t the degree t o which a polymer network w i l l
swell before reaching equilibrium, The f i r s t i s the number and d i s t r i b u t i o n of cross links. The more cross links i n a network, the lower the degree of swelling. Second is t h e molecular weight of the individual chains.
The longer the chains the fewer c m s s l i n k s needed t o bring about a three
dimensional system. Finally a t h i r d important f a c t o r is the chemical
nature of the protein, that is, the type and d i s t r i b u t i o n of R groups along
t h e backbone of the chain.
A s u r f e i t of hydrophobic R groups w i l l decrease the equilibrium
swelling of t h e protein by water whereas a large c o l l e c t i o n of ionizable
(basic o r a c i d i c ) groups i n R w i l l cause the protein t o swell considerably.
When pendant ionizable groups are present, the equilibrium swelling w i l l
incmase a t both high and low pH, but will reach a minimum a t the isoe l e c t r i c point just aa s o l u b i l i t y does f o r soluble proteins,
In summary, the chemical properties of proteins, i n p a r t i c u l a r
their behavior i n acid, base and n e u t r a l aqueous solution are dependent upon t h e constituent amino acids which form the polypeptide chain. The
p r i n c i p a l chemical f e a t u r e s of the peptide chain w i l l be embodied i n t h e
s t r u c t u r e and chemical nature of the pendant R groups. The physical nature
of the peptide chain w i l l depend upon the r e p e t i t i o n of t h e R groups along
the chain. Regularity w i l l encourage the c r y s t a l l i n e state t o form thus
requiring g r e a t e r interactions between the protein and t h e solvent t o overcome t h e c r y s t a l l i n e forces and cause solubilization. I r r e g u l a r sequences
of R will encourage amorphous arrangements and w i l l considerably reduce the
amount of i n t e r a c t i o n necessary f o r dissolution.
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Whcn the R groups a r e joined by chemical cross links, c r y s t a l l i n i t y
i s diminished b u t t h e protein w i l l remain insoluble unless degraded, The
polymer will swell however, because of the interaction between the chain and
the solvent u n t i l the network has stored s u f f i c i e n t energy t o counteract the
solvent-polymer interactions through i t s own e l a s t i c force. The degree of
swelling w i l l be highly dependent on the chemical nature of the R gmups,
the pH of tile solvent and the number of cross links. mrther changes i n
cross linking can occur by the introduction of multivalent ions which would
tend t o f o r a inter-molecular cross links.
Finally, protein s t r u c t u r e is complex i n nature. It has been
shown that some proteins can arrange i n t o mare than one c r y s t a l l i n e form
(allotropy) and t r a n s i t i o n s f r o m one t o t h e other cam be readily obtained by
s u b t l e changes i n p H o r temperature, In addition proteins are subject t o
degradation (denaturization) e i t h e r through cleaving of the main peptide
chain (low probability), i r r e v e r s i b l e rearrangement of c r y s t a l l i n e structure,
disarrangement t o a random s t r u c t u r e , non-reversible dissolution, o r a l t e r a t i o n of pendant chemical groups by heat, ions, and pH changes.
The fundamental causes of changes which take place i n animal muscle
tissue will be snore thoroughly understood once the chemistry and physics of
large molecules has advanced t o a stage where more d e t a i l e d knowledge o f the
charscter of chemical bonding can be detennined more precisely a t the molecular l e v e l*
e w i l l hold the questions f o r the time being, when
MR. PEARSON: W
we come t o t h e final discussion we w i l l a l l o w time f o r the questions from
t h e speakers.
Next t o p i c t h i s afternoon i s t h e Physical Characteristics of
Muscle Tissues as Related t o Imbibed Water. George Wilson, American Meat
I n s t i t u t e has consented t o discuss t h i s ,
MR, GM)RGE WILSON: Thank you, Al. I think a t t h i s point I f e e l
myself i n a position that we Dr. Ibty has, Dr. Schweigert has on occasion
a t t h e Foundation, of explaining t o some of our contributors t h a t we do
have a basic f'undamental program on whether it i s meat hydration o r meat
pigments, o r whatever it might be, t o answer their question as t o has t h i s
got anything t o do with the price of hot dogs, o r t h e p r i c e of mutton. I
t h i n k perhaps it is the sssignment, some of t h e rest of us on t h e program,
I hope others w i l l be able t o come through t o explain a l i t t l e b i t w h a t
e f f e c t s properties of colloids do have on the price of fra.nkCurters o r
some other commodity that we put on the market.