Nonenzymatic Protein Function and Protein Analysis Flashcards
motif
repetitive organization of secondary structural elements together
collagen
has a trihelical fiber; makes up most of the extracellular matrix of connective tissue; strength and flexibility
elastin
extracellular matrix of connective tissue; stretches and recoils, restoring shape to cell
keratin
intermediate filament proteins found in epithelial cells; contribute to the mechanical integrity of the cell and also function as regulatory proteins; make up hair and nails
actin
makes up microfilaments and the thin filaments in myofibrils; most abundant protein in eukaryotic cells; have a + side and a - side, allowing motor proteins to travel unidirectionally along actin filament
tubulin
makes up microtubules; provide structure, chromosome separation and intracellular transport; has + and - side
myosin
primary motor protein that interacts with actin; can be involved in cellular transport
kinesin
align chromosomes; depolarizing microtubules; vesicle transport + end of microtubule
dynein
cilia and flagella; vesicle transport towards - end of microtubule
binding proteins
transport or sequester molecules by binding them
cell adhesion molecules (CAMs)
aid in binding the cell to the extracellular matrix of other cells
cadherin
group of glycoproteins that mediate calcium-dependent cell adhesion
integrins
group of proteins that all have two membrane-spanning chains called alpha and beta; chains are imporrtant in binding to and communicating with extracellular matrix ; role in cellular signaling and can greatly impact cellular function by promoting cell division, apoptosis and other processes
selectins
bind to carbohydrate molecules that project from other cell surfaces
antibodies (immunoglobulins)
proteins produced by B-cells that function to neutralize targets in the body and then recruit other cells to help eliminate the threat
antigen-binding region
within this region are specific polypeptide sequences that will bind on specific antigen sequence
outcomes of antibody binding
- neutralizing the antigen
- marking antigen for destruction by other white blood cells, process known as opsonization
- clumping together (agglutinating) the antigen so it can be phagocytized
ion channels
proteins that create specific pathways for charged molecules
facilitated diffusion
type of passive transport, diffusion of molecules down a concentration gradient through a pore in the membrane created by transmembrane protein
ungated channels
have no gates
voltage-gated channels
regulated by membrane potential change near the channel
ligand-gated ion channels
binding of a specific substance or ligand to the channel causes it to open or close
enzyme-linked receptors
membrane-spanning domain, ligand-binding domain and catalytic domain
G protein coupled receptors
seven membrane spanning alpha helices; integral membrane protein involved in signal transduction
heterotrimeric G protein
have an intracellular link to guanine nucleotides (GDP and GTP)
three main types of G protein
- Gs stimulates adenylate cyclase, increasing cAMP levels
- Gi inhibits adenylate cyclase, decreasing cAMP levels
- Gq activates phospholipase c, which cleaves a phospholipid from the membrane to form pip2. pip2 is then cleaved into DAG and ip3. ip3 can open calcium channels, increasing calcium levels
electrophoresis
subjects compounds to electrical field, which moves them according to their net charge and size
migration velocity
v=Ez/f v=velocity E=electric field strength z=net charge of molecule f=frictional coefficient
polyacrylamide gel
standard medium for electrophoresis; smaller particles pass more quickly
polyacrylamide gel electrophoresis (PAGE)
method for analyzing proteins in their native states; useful to compare the molecular size or charge of proteins
sodium dodecyl sulfate-PAGE (SDS-PAGE)
separates proteins on the basis of relative molecular mass alone; SDS is a detergent that disrupts all non covalent interactions
isoelectric focusing
separating proteins based on their isoelectric point; protein stops moving when the gel pH equals its PI
chromatography
require homogenized protein to be fractionated through a porous matrix;
chromatography process
- place sample on stationary phase or absorbent
- run mobile phase through stationary phase
- retention time is the amount of time the compound stays in the stationary phase
- partitioning is the separation of the compounds based on their varying retention times
Column chromatography
a column is filled with silica or alumina beads as an absorbent and gravity moves the solvent and compounds down the column; less polar compounds move more quickly;
ion-exchange chromatography
solvent beads are charged so a positive charged column will attract and hold negatively charged compounds and vice versa
size-exclusion chromatography
beads in the column have tiny pores of varying sizes; the small compounds go into the holes while the large molecules keep moving
affinity chromatography
a column is coated with a substance that binds a particular protein, retaining it
ways to determine protein structure
x-ray crystallography and nuclear resonance spectroscopy
Edman degradation
uses cleavage to sequence small proteins (50-70aa)
UV spectroscopy
analysis to determine concentration of proteins
bradford protein assay
mixes a protein with blue dye; dye is protonated and green-brown in color props to mixing with proteins; dye binds to proteins, turns blue; higher protein concentration means more blue
