BICD 110 midterm 1

resting potential in animal cell

dependt on potassion ions pass through non gated channels keep potential negative at -70mV

electrical gradient

difference in electrical charges between the inside and outside of the cell

diffusion

diffuse across membrane with no assistance, includes small gasses and small uncharged molecules, travel high to low []

periphenial membrane proteins

do not directly contact the hydrophpbic core of the bilayer, bound to the membrane indirectly by interactign with lipid heads doesnt cross membrane but can be on both sides

F class pump

does not require the phosphylation of proteins to function, opposide to V class, release H down [] grad and is used to power ATP synthesis from ADP and Pi, commonly called ATP synthases

cryo-electron miscroscope allows vizualization of samples without fixing

electron beam to image molecule, uses rapid cooked sample, hold structure

light microscopy

examine cell and tissue, not colloed, have to be fixed and stained, hematoxylin purple stain nuc, Eosin pink for cell

ABC protein functins

export drugs and toxins from cells , can export hdrophobic and hydrophilic molecules, first found in tumor cells

plasma mbm

half proteins half lipids, protiens expressed in plasma membrane dictate function, defines cell shape and encloses cell, controls movement of molecules in and out

transmission EM

heavy metal must be used to prepare the sample in order to add contrast and scatter electron beam, sample absorbed to the EM coated inplastic and carbon, next hevay metal are added megatuve staining occurs, and excludes regions where the sample is adheredto the grid

electron micropscopy

high image resolution, teo types transmission EM and scanning EM

confocal microscopy images

images each focal plane seperatly and excludes light from other out of focus planes

how to keep cell culture alive

imitate in vivo conditions, pH, temp, ionic strength,

cholesterol in phosphatidylcholine

increases thickness of the membrane by making it more ordered Cholesterol increases the thickness of phosphoglyceride bilayers

cytoslic

innerleaflet mbm

basolateral

inside bloodstream, facing side

glucose transport from intestinal lumen to the blood

intesinal lumen(apical) 2na/glucose sympoter (high in gietary glucose, sodium, and cl) moves into cytosol, low na high K, glucose goes through GLUT2 into the blood which has high na low k, blood na/k pump bring already high na into blood and cycles K in and out of cytosol

differential centrifugation

isolate organelles, spin at different speeds and increase speed while decanting supernantent each time to get smaller molecules, can also separate with magnet, increasing dentist top to bottom of separation

stigasterol

main sterol in plant mbm

PE = Phosphatidylethanolamine,

makes curved membrane (small head group)

Peripheral membrane proteins

may noncovalently interact with phospholipid heads becasue they dont make direct contact with the hydrophobic core of the bilayer

most integral proteins have at least one _?

membrane spanning alpha helix

PC = Phosphatidylcholine, most abundant, makes flat sheet-like membrane(big head group)

most abundant, makes flat sheet-like membrane(big head group)

-Gangliosides:

most complex sphingolipids that are abundant in ganglion cells ofnervous tissue

flippase enzyme

move lipids across leaflet, but require ATP, unfavorable

antiporters

move two molecules in opposite directions, three na in push one ca out, cell likes low [] of ca and pushes it out, medications that prevent this stop ca from leaving the heart in pateints with heart faliure since more ca in the cell strengthes heart contractions

signal anchor sequence SA

no cleavble ER signal, positive charge residue near NH3 to make it go in the cytosol which has a negative charg, like foling thread the pushing through end to make c terminus in lumen

cholesterol in sphingomyelin bilayer

no effect on already ordered sphingomyelin bilayer

lysosomes

oftern budded from golgi, contain degradative enzymes that break down molecules into smaller components, acidic lumen pH 4.5 break down proteins

cystic fibrosis transmembrane conductance regulator (CFTR) is an ABC transporter

one function to move cl- down grad certain regulatory region not present in other ABC transporters must be phosphorylated

PS = Phosphatidylserine

only in inner leaflet (cytosolic), marker for apoptosiswhen detected outside cell

N- terminal and internal stop transfer anchor sequence

orient N terminus in lumen and c terminus in cytosol, when STA enter translocon, translocon open like clam and and makes a transmembrane anchor, continuing translation and c terminus remains in cytosol

limitaions of fluorescence microscopy

out of focus fluorescence, thich sections must be imaged separately and then reconstructed

exoplasmic

outter leaflet mbm

Na+/K+ pump

p class pump, moves 3 sodium out and 2 potassium in per ATP hydrolized against concentration, generated low Na high K in cytoplasm , moves sod and pot aginast grad, lead to neg charge in cell,

function of memebrane

pass ions, protect inside, diffusion, cell structure, permeability barrier, control movement of molecules, proteins expressed in mbm dictate cells function, define cell shape

biomembranes three claases of lipids

phosphoglycerides, sphingolipids, sterols

Golgi complex

process and sort secreted lysosomal and membrane proteins on the way to the surface of cell, a stack of flattened membranes called cisternae

scanning electron microscopy

produce 3D image of surface of an unsanctioned sample

Eukaryotes includes

protists, fungi, plants, animals

V class ion pumps

proton pump, does not require the phosphylation of proteins to function. ytansport only protons to generate low pH in plave vacules and animal lysosomes to make acidic , pump against grad into cell

FACS: fluorescence activated cell sorter

purify whole cell, fluorescent tqg and antibody to a cell surface molecule, purify different cells based on proteins expressed

p-pump

require phosphorylation of protiens subunit, at least one of two idential alpha catalytic subunits becomes phosphorylated on a highly conserved aspartae residue ex. H+/K+ pump in stomach H out K in(keep charge in membrane unchanged) Na+/K+ pump

sympoters

same direction ex. nodium move into cell favorable to continue unfavprable pushing glucose inside when it is already highly concetrated inside, also does this with amino acids (two na to 1 aa, glucose, or leucine)

more fluid

shorter chains, unsaturated, high temp,

SRP

signal recognition peptide, bind to ER signal sequence and large ribsomoal subunit and takes this complex to the ER and binds to its receptor on the membrane, SRP receptor, GTP hydrolysis accompanies the disassembly of SRP and its receptor takes unfinished protein chain and ribosome to site on ER membrane where translocation can take place

intestinal cells apical side

specialized in absorbtion of sugars, aa, and other molecules produced by fppd breakdown

topology of a membrane protein

specifies the number of transmembrane segments and their orientation across the membrane

Peroxisomes

spehrical organelle the contain oxidase to detoxify various molecules and break down fatty acids to produce acetyl groups fro biosynthesis

ER

synthesis 33% of protein, largest internal membrane system Rough: surface covered with ribosomes, site of synthesis and secreted proteins, smooth: site of fatty acid and phospholipid synthesis

membrane potential

the electric potential (voltage) across the membrane

why cant v class pumps acidify the lumen by itself

they are lelectrogenic =, the mbm potental will stop more protons from going acoss the membrane by adding negatuve anion cl with each H

immunofluorescence (IF) micoscopy

to visualize specific protiens within cellm cells or tissue chemically fixed, antubiotics applied and dyed,

apical

top surface facing side

porins

transmem proteins that lack and alpha helix, consist of trimers of identical subunits, form barrel with hydrophilic pore, found in membrane of gram-negatuve bacteria such as e coli to protect against harmful agents

channels

transport water, specific ions, or hydrophilic small molecules across the membrane down their gradients, facilitated diffusion/passive transport NO ATP -fast, no confirmational change

Chloroplast

two membrane organelle, thylakoid memebranes conaing enzymes of the photosynthetic pathway, convert light to ATP

Mitochondria

two membranes and generate ATP, 25% of cytoplasm volume, (f): complete the degredation of glucose by oxidation to generate ATP and power cell, folds called cristae, central space called matrix, own set of maternal DNA

transcellular transport

two stage process of the impirt of molecules through the plasma mbm on the apical side and thier export through the basolateral side

transporters consist of

uniporters and cotrasnporters

cotrasnporters

use energy stored in elecrochemical gradient to power transport of molecules, always coupled together, present in all orgs

PI = Phosphatidylinositol,

used in GPI lipid anchored membrane proteins

transporters

variety of ions and molecules, slower than channels, NO ATP

double labeled fluroescence

visualize distribution of two proteins

Fluorescence microscopy

visualize molecules within cell, absorb light at one wavelength and emit at another

inactivation of CFTR

water flows out with na which also follows cl to maintain electrical neutrality, in many CF patitents Phe508 is deleted so build up of mucus outside of cell

During transcellular transport of glucose in an intestinal epithelial cell, which of the following is/are moved against its gradient? Please choose all that apply. A) Import of glucose through Na+/glucose symporter B) Export of glucose through GLUT2 C) Import of sodium through Na+/glucose symporter D) Export of sodium through Na+/K+ pump E) Export of potassium through K+ channel

-Import of glucose through Na+/glucose symporter -Export of sodium through Na+/K+ pump

cell theory

-all living things are composed of one or more cells -every cell comes from a pre-existing cell, -cell is the basic unit of structure, function, and organization in all organisms -cell contains hereditary info which passes cell to cell during division -all cells have the same basic chemical compositions ex. similar set of proteins -all energy flow (metabolic and biochem) of our life occur w/in cell

how to study a cell

-cell culture: grow up mammalian cells, -primary cell cultres: directly from animal tissues or embryos -cell strain: lineage of cells originaling fromone primary culture -cell line: cells undergo oncogenic mutation and continue dividing HeLa(human), COS-7(monkey kidney), CHO (chinese hamster ovary)

what does an alpha helix consist of

20-25 hydrophobic uncharged aa only long enough to span membrane

Where are cholesterols found?

50-90% in plasma mbm and associated vesicles, too hydrophobic to form bilayer so they intercalate between phospholipd molecules to be incorpotaed in mbm, provide structurak support

concentration gradient

A difference in the concentration of a substance on diff sides of the membrane

ABC superfamily

ATP Binding Cassette -diverse function, each protein is specific for a single substrate, four core domains: two transmembrane domains and two cytosolic ATP domains

Although many types of vesicles are similar in size and density, if they are coated with different proteins, it is possible to isolate specific types of vesicles through the use of:

Antibodies attached to bacterial carriers and low speed centrifugation, antibodies will tag the specific vesicles

Type I membrane proteins have all of the following properties EXCEPT

B) Internal signal-anchor sequence

Protein insertion into the mammalian ER membrane typically occurs

B. as translation is happening (cotranslational)

prokayotes include

Bacteria and Archaea

All the following statements describe biomembranes EXCEPT: A. Different biomembranes may contain different proportions of the same phospholipids B. The two leaflets of the same membrane may contain different phospholipids C. Some biomembranes have free edges D. Some phospholipids and cholesterol may cluster to form lipid rafts

C. Some biomembranes have free edges, they do not they are all enclosed

what forms lipid rafts

Cholesterol and sphingolipids

Sterols

Cholesterol is the major animal sterol and is abundant in theplasma membrane of mammalian cells, but is absent from plantand most prokaryotic cells, not phospholipids but amphipathic

Which of the following classes of lipids is (are) present in biomembranes? A. Phosphoglycerides B. Sphingolipids C. Sterols D. All of the above

D. All of the above

As a budding cell biologist, you would like to study your favorite protein, titin, in muscle cells. (Titin is the single largest known protein in a cell, and stretches across a muscle cell sarcomere like a bungee cord during muscle contraction). If you would like to see and be able to distinguish (tell them apart) both titin and the actin filaments that structure the muscle cell, what kind of technique would be best to use?

Double label fluorescence microscopy because you can see the over lap of the two proteins in relation to eachother

region of trans mbm protein

Extracellular (hydrophilic) Membrane-spanning (hydrophobic) Cytosolic (hydrophilic)

Which of the four classes of ATP-powered pumps share overall similarity, including several subunits, the same general organization, and a similar function in being H+ transporters?

F-class pumps and V-class pumps, -F class move H+ down [] grad -v class move H= against [] grad

How are lipid-anchored exoplasmic proteins tethered to the membrane?

GPI anchor

what accompanies SRP disassembly

GTP hydrolysis accompanies the disassembly of SRP and its receptor takes unfinished protein chain and ribosome to site on ER membrane where translocation can take place

How do the following molecules normally travel across a membrane? Glucose up its gradient Glucose down its gradient O2 or CO2 down their gradients K+ down its gradient

Glucose up its gradient- symporter with Na+ Glucose down its gradient -uniporter O2 or CO2 down their gradients-Diffusion K+ down its gradient-channels

Phospholipids with short or unsaturated fatty acyl chains

Increase membrane fluidity, unsaturated=bdl bonds which creakes kinks and more disorder, shorter chains=less LDF so less ordered

The _____________________________ are transmembrane proteins.

Integral membrane proteins

The resting membrane potential in animal cells depends largely on nongated ____ channels.

K+, b/c resting mbm potential is -70mV

ABC superfamily proteins are thought to act as ATP-dependent flippases in transporting:

Lipophilic drugs out of mammalian cells

endosymbiotic theory

Mitochondria and chloroplasts evolved from symbiotic bacteria engulfed though endocytosis by the euk cell evidence: both have two membranes, inner likely from the bacterial membrane, the outer is a vestige of plasma mbm from engulfment, broth have own GNA genome, synthesis of proteins in these more closely related to bacteria

ATP powered pumps

Move specific ions or molecules across the membrane; require ATP molecules to function, against a gradient, active trasnport, slowest transfer b/c of confirmational change, all have binding site for ATP in cytosolic region of protein

If a cellular homogenate were subjected to differential centrifugation, which of the following would be expected to pellet first?

Nuclei, because 1/2 cell size, very large

Types of ATP powered pumps

P-class, V-class, F-class, ABC superfamily,

phosphoglycerides

PE = Phosphatidylethanolamine, makes curved membrane (small head group) PC = Phosphatidylcholine, most abundant, makes flat sheet-like membrane(big head group) PS = Phosphatidylserine, only in inner leaflet (cytosolic), marker for apoptosiswhen detected outside cell PI = Phosphatidylinositol, used in GPI lipid anchored membrane proteins

Prokaryote vs. Eukaryote cells

Prokaryotes:.5-1 micron, no nucleus, free-flowing single c circular chromosome eukaryotes: 10 micron, nucleus, mbm bound organelles, Both: ribosomes, transcription/translation, , proteins and ATP, ETC, plasma membrane, cell walls, flagella

cotranslational translocation

Simultaneous transport of a secretory protein into the endoplasmic reticulum as the nascent protein is still bound to the ribosome and being elongated.

Integral membrane proteins (Transmembrane

Span a phospholipid bilayer and consist of three segments- Have a helix or b sheet domain, some are receptors, not all transport material

Sphingolipids

Sphingomyelin is themost abundant -Sphingomyelin: Sphingomyelins are present abundantly in the cell membranes of animal cells especially in the myelin sheath of neurons -Glycosphingolipids: abundant in the brain and spinal cord (Glucosylceramide,(GlcCer) shown above) -Gangliosides: most complex sphingolipids that are abundant in ganglion cells ofnervous tissue

-Sphingomyelin

Sphingomyelins are present abundantly in the cell membranes of animal cells especially in the myelin sheath of neurons

electrochemical gradient

The diffusion gradient of an ion, which is affected by both the concentration difference of an ion across a membrane (a chemical force) and the ion's tendency to move relative to the membrane potential (an electrical force).

Ergosterol

The main sterol in fungal membranes

translocon

a complex of proteins forms a channel through ER membrane, continues the elongated protein chain into the central pore of the translocon

-Glycosphingolipids

abundant in the brain and spinal cord (Glucosylceramide,(GlcCer) shown above)

cytosolic side acylation (Lipid-anchored membrane proteins)

acylation, anchored fatty acyl group cov attacheed to N terminal glycine reside image: cytosolic leaflet with gly attached to potein chain with COO- in cytosol

why do integral proteins-spanning membrane have alpha helixes

alpha helixes are stably embedded becasue of energetically favorable interactions, single alpha helix suffiencent to wmbed protein,

charged residues in alpha helix guide assembly of multimeric proteins

also contain charged residues, guide assembly and stabilization of multimeric proteins

Lipid-anchored membrane proteins: GPI anchor

always have phosphatidylinositol, with sugar residues attached on top, with phosphoethanolamine attached above sugar holding protein

Phosholipids

amphopathic lipid with a phophate base head group and a two pronged hydrophobic tail

prenylation (Lipid-anchored membrane proteins)

anchored to membrane by hydrocabon chain attached to cysterine residue at or near the C-terminus, NH3 in cytosol

cholesterol

animal

method to obtain proteins of intrest

anitbodies, monoclonal antibodies generated in a single cell type, polyclonal antibodies generated multiple cell types, inject proteins of interest into animal and let antibodies against protein epitope(part of antigen the antibody attach to) be raised, can tag with magnet or fluorescent

A characteristic of all biomembranes is

asymmetry, Cholesterol, however, seems to be evenly distributed• When cells die, the asymmetry is no longermaintained and lipids normally enriched in thecytoplasmic leaflet can be detected in the exoplasmicone

N terminal ER signal sequence

at N terminus of protein, stretch of hydrophobic amino acids and removed by signal peptidase., not present in mature form of protein, 16-3o hydrophobic residues, all signal scontain one or more postively charges aa

transcriptional fusion

attach promoter and gfp, promoter express gfp and flurences shows what cells this promoter is activated

translational fusion

attach promoter with gene and gfp so protein is trascribed with gfp and follows protein

multipass proteins

both SA and STA alternating, two or more membrane spanning segments=helices

Lipid-anchored membrane proteins

bound covalently to lipid molecule, hydrophobic segment of the lipid embed into mbm and anchors protein

unitransporters

carrier single molecule or ion unidirectionally, down gradient, ex. glucose into cell, trasnporter called GLUT1

less fluid

cholesterol, long chains, low temp, sphingolipids

chemical gradient

concentration gradient for an ion across the plasma membrane

Nucleous

contains DNA of genome, half size of cell, site of transcription, mRNA leaves through nuclear pores, two membranes the outer membrane is continuous with rough ER