2. Protein Structure and Function

Lecture 2

which amino acid structure determines protein folding and therefore protein function

the primary structure (amino acid sequence)

how can proteins lead to disease

if they are improperly folded, defective, or absent

what roles can proteins perform

enzymes, structural components, transporters, channels, receptors, motor proteins, and some hormones

Protein structure and conformation summary

DNA determines amino acid sequence → amino acid sequence determines protein structure → protein structure determines protein function

What holds a protein together

H-bonds, Ionic bonds, Hydrophobic interaction, Van der Waals forces, and Disulfide bonds

what makes a protein work?

the 3D conformation and the forces holding the protein together allows it to bind ligands, substrates and other proteins in highly specific ways

Protein functioning and stability

proteins are stable not because of one strong bond, but because of thousands of weak interactions acting cooperatively

what is the native conformation of a protein

the highly organized 3D pattern of folding to make a protein active; lowest free-energy

protein conformation definition

the 3D pattern of folding to make a protein function

causes of protein denaturation (unfolding)

heat (disrupts H-bonds and stabilizing interactions causing unfolding and loss of function)

changes in pH (interfere with hydrophobic interactions, expose hydrophobic residues in protein core, promote unfolding)

toxins and heavy metals (bind cysteine residues and other functional groups altering structure and inhibiting enzymatic activity, potentially leading to irreversible denaturation)

clinical relevance of protein stability/denaturation

because proteins operate within a narrow physiological range of temperature, pH, and chemical conditions, relatively small environmental changes can impair protein function and contribute to disease

protein rigity

proteins are not rigid structures. they are dynamic molecules that undergo small, reversible conformational changes as part of their normal function

are changes in proteins shape/structure always harmful

no, conformational changes can be used to regulate protein activity and function

phosphorylation

the addition of a phosphate group. when added to a protein, it changes conformation and laters activity. Usually serine, threonine and tyrosine are targets of phosphorylation. can activate or deactivate the proteins function.

examples of modifications to alter protein structure

phosphorylation, methylation, ubiquitination, acetylation

what can PTMs affect besides activity

stability, localization and interactions with other molecules

allosteric regulation

binding of a ligand at allosteric sites induces a conformational change that can increase or decrease the proteins activation

what is the induced fit model

a conformational change in an enzyme triggered by substrate binding that improves catalysis, which can help facilitate product release

protein conformation and stability summary

proteins function because they are dynamic molecules. the same weak non-covalent interactions that stabilize protein structure also allow proteins to undergo regulated conformational changes that control biological activity

Protein structure is not static

regulated conformational changes are often the mechanism by which proteins perform their biological functions. conformational changes are thus integral to the regulation of protein function, allowing proteins to respond dynamically to various cellular signals and conditions, ensuring precise control of biological processes

key point of protein conformation

proteins fold into specific conformations to function. this 3D conformation is primarily stabilized from noncovalent bonds. noncovalent bonds are relatively weak and can easily be broken by changes to the local environment. some changes in the local environment (+ or - phosphate bond) are used advantageously to control the activity of a protein

the primary structure of a protein

the linear sequence of amino acids linked by peptide bonds that determines its conformation and function

mutations in protein primary structure clinical correlation

mutations can alter the amino acid sequence. depending on where the change occurs, the mutation may have little effect, reduce protein function, create new function, or disrupt folding altogether. some are beneficial while others are harmful

sickle cell disease results from a single amino acid substitution demonstrating how a small change in primary structure can dramatically alter protein structure and function

gene duplicaiton

during evolution, genes may become duplicated within the genome. once duplicated, the extra can accumulate mutations and eventually evolve a new or specialized function

example of protein family that arose through repeated gene duplication and divergence

keratin proteins

isoforms

related proteins with similar functions but slightly different amino acid sequences. These differences often optimize protein function for particular tissues or developmental stages

isozymes

enzyme isoforms that catalyze the same reaction bit differ in regulation or tissue distribution

secondary protein structure

localized, repetitive folding patterns of the polypeptide chain. stabilized by H-bonds. forms a-helix, B-sheet/B-strandswh

why are there 2 types of secondary structure

bulky R groups can only allow certain angles of rotation around the a-carbon, or charge attraction and repulsion can limit the range of available angles

driving factor of secondary protein structure summary

secondary structure is driven by backbone H-bonds and constrained by steric limitations of the polypeptide backbone and side chains

Supersecondary Protein Structure

supersecondary structures are recurring, stable arrangements of 2 or more elements of secondary structure known as a protein motif

motifs are intermediates between secondary and tertiary structure

often associated with specific structural or functional roles

unlike protein domains, motifs do not typically function independently but contribute to the overall tertiary structure and function of the protein

supersecondary structure summary

protein motifs are recurring combinations of secondary structures that support specific structural or functional roles within proteins

Tertiary protein structure

overall 3D folding of a single polypeptide chain. can be considered the conformation of a polypeptide

determined by primary structure and held together by noncovalent forces and disulfide bonds

discussed in broad groups (fibrous proteins, globular proteins, and membrane proteins)

protein domains

larger, functionally and structurally independent units of a protein that often represent modular units of protein function and evolution.

often serve as modular functional units

many proteins contain multiple domains with distinct functions

domains are frequently conserved throughout evolution

a Kringle Domain

type of protein structural domain characterized by a looped, triple-disulfide-linked structure. it is named for scandinavian pastry. these domains are important in protein-protein interactions, particularly in the blood coagulation and fibrinolytic systems.

Quaternary structure

functional protein complex comprised of 2 or more interacting polypeptide subunits; also a complex of multiple polypeptides or subunits held together by various interactions

stabilization of quaternary structure

held together primarily by hydrogen bonds, ionic interactions, hydrophobic interactions, Van der Waals forces

some extracellular proteins can be additionally stabilized by disulfide bonds

Advantages of multi-subunit proteins

binding of a molecule to one subunit can influence the activity of other subunits.

different subunits may serve different functions.

multienzyme complexes can transfer intermediates directly between enzymes (substrate channeling)

Quaternary structure summary

allows proteins to coordinate activity between multiple subunits, enabling regulation, cooperativity, and functional specialization

protein folding and conformation errors/consequences

alterations in folding can impair activity and contribute to disease

rotation of the polypeptide backbone and restrictions

occurs mainly around the phi and psi bonds.

due to the partial double bond character of the peptide bond, rotation is restricted around the bond itself

interactions between amino acid side chains can also restrict rotation, and determine the folding pattern

role of side chains in protein folding

the size, charge, polarity, and chemical properties of the dise chains strongly influence folding.

nonpolar residues in core, polar and shared located on the protein surface where they interact with water

protein stability and denaturation

because proteins function optimally within narrow physiological conditions, changes in temperature, pH, or chemical environment may lead to loss of function and disease

hierarchical model of protein folding

localized folding into regions of secondary structures

longer-ranges interactions then occur between these regions

motifs and domains form

protein ultimately reaches its native conformation

this model emphasizes progressive and structural organization

molten globule model

hydrophobic collapse

hydrophobic residues rapidly cluster away from water

secondary structure is present, but side chains are not yet tightly packed into their final positions

this model emphasizes early collapse driven by hydrophobic interactions

thermodynamic funnel model

incorporates features of both models

proteins can follow many possible folding pathways

at the top of the funnel, the unfolded protein has many possible conformations and high entropy

as folding proceeds, stabilizing noncovalent interactions form

entropy decreases and free energy decreases

the native conformation lies at the bottom of the funnel because it represents the most thermodynamically stable state

why are hydrophobic residues usually buried in the protein core or in the lipid spanning portion of membrane proteins

hydrophobic amino acids tend to cluster in the interior away from water. they are buried away from water because the arrangement is energetically favorable

in membrane proteins, they’re commonly found in membrane spanning regions where they interact with the hydrophobic lipid bilayer

where are polar residues usually in regards to the protein

on the surface or in clefts/crevices that are in contact with water or substratesp

protein conformation and folding summary points

heavily influenced by the chemical properties of amino acid side chains and the environment surrounding the protein

hydrophobic residues tend to avoid water, while polar residues interact favorably with aqueous environments

protein folding minimizes free energy by optimizing hydrophobic and hydrophilic interactions

Levinthal’s Paradox

highlights the apparent contradiction between the vast number of possible protein conformations and the relatively fast time it takes for a protein to fold into its correct, functional state

highlights the fact that proteins fold far too quickly to do so by randomly sampling every possible conformation

proposed that if a protein were to fold by randomly sampling all possible conformations of its polypeptide chain to find the native state, it would take an astronomically long time to fold, yet proteins typically fold within microseconds to seconds

Levinthal’s Paradox summary points

protein folding is not random

folding proceeds through energetically favorable pathways that progressively stabilize the structure

protein folding follows a funnel shaped energy landscape where the native structure represents the lowest free energy state

proteins fold through intermediate states and preferred pathways, not by random chance

chaperone proteins and the physical properties of amino acids help guide folding efficiently

molten globule model ?

im confused. says the same thing as the previous info?? ask about this

therrmodynamic funnel model?

also saying the same thing as the previous

low free energy state

Chaperones

proteins that assist in proper protein folding without becoming part of the final structure

prevent aggregation of denatured proteins

refold misfolded or partially denatured proteins

keep proteins unfolded for translocation across membranes

heat shock proteins

molecular chaperones that assist protein folding under normal conditions and are upregulated during cellular stress.

stress proteins that aid in protein remodeling when cells are exposed to stress such as high temps or UV irradiation

ex: Hsp70 - binds exposed hydrophobic regions of newly synthesized or partially unfolded proteins

GroEL-GroES

bacterial chaperonins; eukaryotic cells contain related chaperonin systems

provide an isolated chamber where proteins can fold in an ATP-dependent manner

isomerases

rearrange atoms in a molecule to change its structure

Protein disulfide isomerase (PDI)

in the ER

reduced PDI catalyzes the reduction of mis-paired thiol groups (inaccurate disulfide bond formation. catalyzes disulfide exchange reactions that break and rearrange these incorrect bonds until the proper native disulfide pattern is achieved

catalyzes post translational disulfide exchange and aids protein folding by cleaving and reforming disulfide bonds

Peptide prolyl cis-trans isomerase (PPI)

peptide bonds involving proline can exist in either cis or trans conformations

interconversion between these forms occurs slowly and may limit the rate of protein folding. they catalyze this rearrangement and accelerate proper protein folding