Biochemistry Exam 2 (Protein Function)

Main Protein Functions

Structural – example is the cytoskeletal proteins

Enzymatic – catalyze some chemistry

Binding (Transport, Storage) – function by binding to something

Enzymes and Binding Proteins

Function by the reversible binding of other molecules called ligands

Ligands

• bind in a binding site

• Ligands that bind to binding proteins are still called ligands

binding site

a site on the protein where the ligand binds

Substrate

Ligands that bind to enzymes

catalytic/active site

The binding site on the enzyme

Ligan binding Models

• Lock and Key

• Induced Fit

Lock and Key Model

The active site has a fixed, rigid geometrical shape. Only a substrate with a matching shape can fit into it.

Induced Fit Model

Initial binding of the ligand to the protein induces a conformational change in the protein, which causes tighter binding to the ligand.

Myoglobin

• 153 residues

• 8 a-helical segments (~78% of the residues)

• Binds O2 with the Heme prosthetic group

Protoporphyrin

The complex organic ring structure found in several different proteins including myoglobin and hemoglobin

Protein-Ligand Interactions

• Can be expressed in terms of an association constant or a dissociation constant.

• leads to an expression for the fraction of ligand-binding sites occupied by ligand, Y.

Hemoglobin

• an allosteric protein

• binds O2 Cooperatively (positive or negative)

The Bohr Effect

the affect changes in pH has on O2 binding to hemoglobin (high pH= high binding???)

BPG (2,3-Bisphosphoglycerate)

reduces the affinity of hemoglobin for O2 by stabilizing the T-state

T-state

• low-affinity, deoxyhemoglobin form of hemoglobin that stabilizes the release of oxygen

• Low pH, High CO2, high 2,3-BPG

• Facilitates the release of oxygen to tissues

• Resists oxygenation due to hydrogen and ionic bonds

R-state

• high-affinity, oxygenated form of hemoglobin that facilitates oxygen binding

• High pH, low CO2, low 2,3-BPG

• Facilitates the binding of oxygen in the lungs

• Oxygen binding causes a conformational change, allowing more oxygen to bind to other subunits

immune response

A coordinated set of interactions among many classes of proteins, compounds (molecules), and cell types

• distinguishes molecular “self” from “nonself” and destroys “nonself”

• eliminates viruses, bacteria, and other pathogens and molecules

leukocytes

white blood cells, including macrophages and lymphocytes

Systems of the immune response

• humoral immune system

• cellular immune system

humoral immune system

directed at bacterial infections and extracellular viruses

• uses antibodies produced by B lymphocytes or B cells

cellular immune system

destroys s T-lymphocytes, Helper T-cells, and memory cells

antibodies

immunoglobulins (Ig) = bind bacteria, viruses, or large molecules identified as foreign and target them for destruction

• produced by B lymphocytes or B cells

T lymphocytes (cytotoxic T cells (T_C cells))

recognition of infected cells or parasites, involves T- cell receptors on the surface of T_C cells

helper T cells (T_H cells)

produce soluble signaling proteins called cytokines

• interact with macrophages

• stimulate the selective proliferation of T_C and B cells that can bind to a particular antigen (clonal selection)

memory cells

permit a rapid response to pathogens previously encountered

antigen

molecule or pathogen capable of eliciting an immune response

• can be a virus, a bacterial cell wall, or an individual protein or other macromolecule

• antibodies or T-cell receptors bind to an antigenic determinant or epitope within the antigen

• antigenic determinant/epitope: the part of an antigen that is recognized by the immune system

haptens

small molecules that can elicit an immune response when covalently attached to large proteins

immunoglobulin G (IgG)

major class of antibodies

• one of the most abundant blood serum proteins

• 4 polypeptide chains: 2 heavy chains and 2 light chains

• cleavage with protease papain releases the basal fragment Fc and two Fab branches (each with a single antigen-binding site)

• constant domains contain the immunoglobulin fold structural motif

The Variable Domains of Immunoglobulin G

Heavy and light chains each have one

• associate to create the antigen-binding site

• allows formation of an antigen-antibody complex

Phagocytosis of Antibody-Bound Viruses by Macrophages

When Fc receptors bind an antibody pathogen complex, macrophages engulf the complex

polyclonal antibodies

• produced by injecting a protein, eg., into an animal

• contain a mixture of antibodies that recognize different parts of the protein

monoclonal antibodies

• Synthesized by a population of identical B cells (a clone)

• homogeneous, all recognizing the same epitope

Primary Motor Proteins

• Myosin and Actin

• Together make up more than 80% of the protein mass of muscle. Involved in ATP-driven conformational changes that result in muscle contraction

Myosin

• 2 heavy chains and 4 light chains

• forms a fibrous, left-handed coiled coil domain (tail) and a large globular domain (head)

actin

monomeric G-actin (Mr 42,000) associates to form a long polymer called F-actin

thick filaments

rodlike structures of aggregated myosin

thin filaments

F-actin along with the proteins troponin and tropomyosin

• assemble as successive monomeric actin molecules add to one end

• each monomer binds and hydrolyzes ATP

Components of Muscle

• each actin monomer in the thin filament binds to one myosin head group

• Additional Proteins Organize the Thin and Thick Filaments into Ordered Structures

muscle fiber

• large, single, elongated, multinuclear cell

• each muscle fiber contains ~1,000 myofibrils, each consisting of thick and thin filaments and surrounded by sarcoplasmic reticulum

The Structure of a Sarcomere

• A band

• I band

• Z disk

• M line

A band

stretches the length of the thick filament

I band

contains only thin filaments

Z disk

attachment site for thin filaments

M line

bisects the A band

How Myosin Thick Filaments interact with Actin Thin Filaments

Myosin Thick Filaments Slide along Actin Thin Filaments

• myosin binds actin tightly when ATP is not bound to myosin

• series of conformational changes due to binding, hydrolysis, and release of ATP and ADP causes muscle contraction

tropomyosin

binds to the thin filament and blocks the myosin-binding sites

Troponin

binds Ca2+ released from the sarcoplasmic reticulum, causes a conformational change, and exposes myosin-binding sites