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MMBIO Quiz 3
| Term | Definition |
|---|---|
| protein domain | region of a protein that has a specific function, location, or shape |
| transmembrane domain | part of the protein that anchors in the lipid bilayer |
| in a transmembrane domain, the ______ amino acids stay inside the lipid tails | hydrophobic this means the hydrophilic domain part is outside, touching the water |
| ion channels | allows ions to travel through membrane |
| nucleosome | basic unit of DNA packaging in cells (so it fits in the nucleus) |
| nucleosomes have DNA wrapped around a core of | histone proteins |
| the nucleosome and histone proteins play a huge role in _____________________ because DNA is wrapped tightly and is harder to access | regulating gene expression |
| helicase | enzyme that unwinds DNA |
| we can modify peptides to _____ or _______ proteins | activate or de-activate |
| all modifications are also | reversible |
| phosphorylation | serine, tyrosine, threonine phosphate added onto a OH (hydroxyl) group |
| proteins dramatically change shape when a ______________ is changed, which is why adding or taking away charged groups changes the function | a charge on a single amino acid |
| acetylation | lysine adding an acetyl group (COCH3) to the amino group (NH3) |
| methylation | lysine adding a methyl group (CH3) to the amino group (NH3) |
| acetylation neutralizes the positive charge on lysine, reducing the attraction between positive histones and negative DNA, which ____ the DNA | loosens |
| methylation blocks the acetyl group from binding which | tightens back up DNA |
| glycosylation | asparagine (N-linked) or serine/threonine (O-linked) addition of sugars folding, stability, signaling, protects from degradation |
| anything that ends with ase is a | enzyme |
| enzyme | protein that catalyzes a chemical reaction, increasing the rate of reaction |
| enzymes work with/on ______ to make ______ | enzymes work with/on substrates to make products |
| lock and key | perfect fit, enzyme shape is constant |
| induced fit | enzyme and substrate change a bit to fit together |
| in a radioactive assay you can label the _____ with radioactivity and after the enzyme converts it to a product, the isotope can be detected | substrate |
| enzymes can ____ something big into something small | cleave, it does this by breaking chemical bonds |
| isolating and separating DNA steps | 1. break open cell membrane, release DNA 2. digest RNA with RNase 3. separate DNA from proteins by phenol extraction 4. precipitate DNA with salt and alcohol, centrifuge 5. dry out DNA 6. dissolve DNA is slightly basic buffer (8-8.5) |
| why cut up RNA with RNase? | easier to destroy it all then separate the two, since they are similar |
| phenol extraction | separating DNA and protein DNA/RNA go to top (hydrophilic, aqueous layer) protein stays in bottom (interface/organic layers) |
| alcohol precipitation | DNA/RNA are highly soluble in water (polar) alcohol less polar add salt, Na+ forms bonds with DNA/RNA as concentration of alcohol gets high DNA forms insoluble salt |
| denaturing a protein | unfolding, stopping them from working |
| phenol extraction _____ proteins | denatures |
| RNA isolation | very similar freeze sample and grind denature proteins separate by centrifugation ethanol precipitate |
| these are old methods, new methods are like | ion exchange chromatography |
| phenol/chloroform is | hazardous new ways are faster and less wasteful (but more expensive) |
| DNA absorbs light at a wavelength of | 260 nm |
| gel electrophoresis | separates DNA by size using an electric current |
| DNA is negative so it moves through the gel to the | positive end |
| smaller pieces move ____ and ______ | faster and further |
| ethidium bromide | loading dye to visualize DNA better in gel is heavy so makes the DNA fall into wells |
| rules of electrophoresis 1 | run a mass ladder (DNA fragments of known size) every time |
| rules of electrophoresis 2 | smallest DNA fragments run fastest |
| rules of electrophoresis 3 | we show gels with smallest fragments at the bottom |
| rules of electrophoresis 4 | brighter band = more DNA |
| rules of electrophoresis 5 | larger fragments are brighter cause more binding sites to dye |
| DNA size | how many base pairs (500 bp vs 2000 bp) affects how far the DNA travels |
| DNA amount | how many copies of the fragment are present 10 copies of a 500 bp fragment or 1000 copies of 500 bp affects how bright the band is |
| agarose gel | lower power, cheaper, faster fragments of similar size are shown as a single bright band |
| polyacrylamide gel | resolve DNA that differ in size by one nucleotide |
| PAGE | separates proteins based on size, shape, and charge |
| SDS | chemical in PAGE that makes all proteins negative |
| BME | chemical in PAGE that breaks apart tertiary and quaternary structures to look at individual peptides |
| plasmids | small accessory DNA molecules in bacteria circular non-essential genes |
| endonucleases | cut in the middle of molecules, cut specific sequences cuts right through |
| exonucleases | cuts at ends of molecules, random takes nucleotides off |
| restriction endonucleases/enzymes | proteins that cut DNA at specific sequences |
| double digest | cut DNA with one restriction enzyme, then cut with a second one fragment remians the same as the first cut - not cut by secnond enzyme fragment changes after second cut - it was cut by second enzyme |
| restriction enzymes are | palindromes (matches up and reads the same back and forward) AGCT TCGA |
| we clone DNA ______ of a promoter so gene will be expressed | downstream |
| sticky ends vs blunt ends | sticky ends - puzzle pieces, overhangs (EcoR1) blunt ends - blocks (Pvull) |
| DNA ligase | joining pieces of DNA together |
| why clone DNA? | simple way to generate a large amount of DNA, which we need a large amount to study |
| why clone DNA? | provides a way to manipulate DNA (express genes, change DNA to learn tits function - mutagenize) |
| cloning vector | a DNA molecule used to carry a DNA inset into a host cell |
| selectable marker | uses natural selection to force bacteria to maintain DNA (antibiotic resistance, so only host cells with vector will survive) |
| bacterial origin | allows vector to replicate independently inside the host |
| multiple cloning sites (MCS) | many restriction enzyme cut sites, makes it easy to insert |
| B-galatosidase reporter gene (optional) | tests plasmids to see if gene was actually inserted blue: no DNA inserted white: gene was inserted, check if it was correct |
| promoter and enhancer element (optional) | expressing a protein, increasing expression |
| process of cloning | combine DNA with vector, ligase put ligated DNA in bacteria spread bacteria on antibiotic plate only those with plasmid will survive grow/culture/duplicate, extract plasmid |
Created by:
anyasalmon