Week 2, Chapters 4 and 5
Week 2, Chapters 4 and 5 BIOL 3510
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This 4 page Class Notes was uploaded by Marin Young on Tuesday February 2, 2016. The Class Notes belongs to BIOL 3510 at University of North Texas taught by Dr. Chapman in Spring 2016. Since its upload, it has received 43 views. For similar materials see Cell Biology in Biological Sciences at University of North Texas.
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Date Created: 02/02/16
Week 2: Chapters 4, 5 | BIOL 3510 Notes by Marin Young Protein Structure and Function: • Proteins' functions rely on their specific structures ○ A key example is binding sites, where a protein grabs onto a ligand (a moleculethat interacts with a protein, like a nucleotide or hormone) The shape, size, and chemistryof the binding site are a perfect fit for the shape, size, and chemistry of the ligand--binding is very specific Non-covalentforces like hydrogen bonds and hydrophobic (Van der Waals, although this is technically a bit of a misnomer)interactions hold ligand to protein These are the same forces responsible for tertiary structure: binding a ligand can change a protein's conformation,or shape ○ Proteins are suited to their environments: a transmembraneprotein might have several hydrophobic alpha-helices in the middle of the plasma membrane and a hydrophilic domain protruding into the cell • Enzymes = proteins that catalyze (facilitate) reactions without being consumed ○ Enzymes lower activation energy--energy needed to start a reaction ○ The thermodynamics(heat change and energy cost) do NOT change ○ Enzymes can help reactions by holding two substrates (reactants) close together, temporarilylending or borrowing electrons, or changing bond angles to stabilize a transition state (hold a substrate in the position needed for bonds to break and form) ○ Enzyme kinetics: reactions go faster when more substrate is available, but still limited by amount of enzyme Theoretically,if there's a huge excess of substrate, everyenzyme molecule could catalyze a reaction, spit out products, and immediatelygrab more substrate--this maximum speed is called v max In reality, the more substrate is present, the less the reaction rate will increase--if almost all enzyme moleculesare "busy," it takes much more substrate to "find" the remaining enzyme molecules Every enzyme has a certain substrate concentration,called K ,mwhere half the enzyme moleculesare bound to substrate □ If the substrate concentrationis morethan K ,mspeed is morethan half v maxand will increase slowly ○ Enzymes can be regulated by feedback loops or regulatory molecules:a product or messenger can influence an enzyme's activity This works because binding a ligand can change a protein's conformation,so binding an inhibitor can switch an enzyme into an inactive conformation □ Negative feedback: the product of a reaction inhibits the enzyme responsible for that reaction (example:pyrimidine synthesis) Allosteric regulation involves specific regulatory binding sites on enzymes, separate from the active site where substrates bind (example: CTP inhibits ATCase) ○ Other ways to modify enzymes: Phosphorylation(adding a phosphate group via a kinase) changes protein conformationdramatically □ Example: mitosispromoting factor (MPF) is phosphorylated twice and dephosphorylated (using a phosphatase) once to becomeactive G proteins becomeactive when they bind GTP and inactive when they hydrolyze it □ Example: EF-Tu can bind tRNA when holding GTP; it hydrolyzes GTP and releases tRNA □ This is also important in signal transduction ATP hydrolysis causes a big conformationalchange called the power stroke in motor proteins that allows them to "walk" along cytoskeletalfilaments □ This happens in myosin in muscle cells: myosin walks along actin to pull the ends of a sarcomere(muscle fiber unit) together and contract the muscle • Technique: antibodies bind very specifically to proteins and can help us study protein levels and activity ○ Injecting a lab animal with an antigen (something that stimulates the immune system, in this case a protein) several times causes the animal to produce lots of antibodies against it These antibodies can be isolated and added to a mixture of proteins--they will bind to the specific protein ○ These antibodies can be isolated and added to a mixture of proteins--they will bind to the specific protein being studied and cause it to precipitate (like agglutination in blood typing), which isolates the protein DNA and Chromosomes: • DNA structure is very hierarchical: there are many levels of organization, from a single nucleotide to an entire chromosome, with their own structures to know • One nucleotide consists of a nitrogenous base, a 5-carbon sugar, and a phosphate group ○ DNA's four bases are adenine, guanine, cytosine,and thymine, and the sugar is deoxyribose ○ Cytosine and thymine are both pyrimidines,which means their structures are based on a six-atom,nitrogen- containing ring ○ Adenine and guanine are both purines, which means they have bicyclic ring structures ("PUGA-2": purines are G and A, with 2 rings) • Nucleotides form base pairs using hydrogen bonds ○ Cytosine and guanine share 3 hydrogen bonds ○ Adenine and thymine share 2 hydrogen bonds ○ Notice that each base pair (bp) has one purine and one pyrimidine--this means each bp is the same width ○ DNA molecules with "high GC content" (more G-C bps than A-T bps) are more strongly bound and have higher melting points due to the higher number of hydrogen bonds • A DNA double helix has two antiparallel strands in a right-handed spiral with 10 bp per turn ○ Antiparallel: one goes 5' to 3' (has a phosphate at one end and a sugar at the other) and one goes 3' to 5' (sugar at one end, phosphate at other) ○ Minor and major groovesalternate because of the offset between the two strands Proteins to bind to DNA based on this pattern/shape • DNA wraps around specialized proteins called histones ○ A histone complex is an octamer with two each of H2A, H2B, H3, and H4 ○ A 147-bp length of DNA wraps twice around each histone complex,with a 50-ish-bp segment of linker DNA between histone complexes ○ Nucleosome= one histone complexplus its wrapped and linker DNA ○ If this "beads on a string" chromatin (DNA + histones) was exposed to a nuclease, the linker segments would be destroyed and yield nucleosomecore particles (histone complexeswith wrapped DNA) ○ Histone proteins contain many Arg and Lys residues (+ charges attract DNA) and are highly conserved among species H3 subunits' tails are most used for regulation • Nucleosomesare packed into 30 nm fibers ○ Histone H1 (NOT part of the octamer)binds between wrapped and linker DNA to hold nucleosomesclose together ○ Histone tails can also interact • 30 nm fibers form 300 nm loops • 300 nm loops are squished and folded together to form a mitotic (condensed) chromosome Epigenetics and Gene Expression: • Regulating gene expression often depends on nucleosomestructure and function Epigenetics and Gene Expression: • Regulating gene expression often depends on nucleosomestructure and function • DNA on nucleosomesnaturally unwraps and very quickly rewraps every so often (so it's unwrapped for 10-50 ms every 250 ms) ○ Sequence-specific binding proteins recognize and bind to a short DNA sequence to stabilize it: these can act fast to bind to unwrapped DNA and let it get transcribed ○ Chromatin remodeling complexes use ATP to slide DNA over a nucleosome, which exposes DNA normallywrapped around the histone complex • Since gene expression requires transcription of the original genome,looser packing of chromatin allows more gene expression ○ Euchromatin ("true chromatin") is looser and easier to transcribe (like the accessible tops of 300 nm loops) ○ Heterochromatin ("other chromatin") is highly condensed (like in a mitotic chromosome) Usually gene-poor regions ("junk DNA") Barr bodies (see box) Helps adjust gene dosage to account for polyploidy in plants ○ Euchromatin and heterochromatindescribe a spectrum of how free or packed chromatin is--it's not one or the other ○ Methylation, phosphorylation, and acetylation are all ways of covalentlymodifying histone tails to adjust chromatin packing Methylationneutralizes a positive charge (often on Lys-9 in the H3 subunit) and causes tighter packing and less gene expression Acetylationand phosphorylation both increase negative charges, which repel negatively charged DNA to cause looser packing and more gene expression ○ Ubiquitin-tagging also modifies chromatin by marking histones for degradation (destruction and recycling) • Patterns of condensation can be replicated with DNA: epigenetic inheritance ○ Enzymes that modify chromatin are present in the cell and modify daughter DNA molecules after replication ○ Stress, like infection with a pathogen (in a plant), can induce epigenetic changes A bit like mutation: genetic variation improveschances that some offspring will survive a rough time period (offspring can becomeresistant to pathogen) History and Biotechnology: • Human Genome Projectwas drafted in 2001, finished in 2004 ○ The human genome has 3.2 billion bps, with about 30,000genes on the 22 autosomesand 2 sex chromosomes ○ About 1.5% of the genomeis made of exons (regions that will be transcribed and translated into proteins) ○ Much of the rest (introns and "junk DNA") helps with structural and regulatory functions (coveredmore in chapter 7) This is important partly because many cancers come from loss of regulation, like a mutation to the p53 gene, which normally monitors DNA and stops division of faulty cells ○ The ends of chromosomeshave repetitivecaps called telomeresthat shorten with everycell division DNA polymerasecan't replicate the "first" few bases because there's no upstream DNA to bind to-- it has to start a few bases into the chromosome DNA polymerasealso can't replicate the "last" few bases on a chromosomefor similar reasons--it would basically fall off That said, rememberchromosomeshave many replication bubbles/origins of replication, not just That said, rememberchromosomeshave many replication bubbles/origins of replication, not just replication from one end to the other • A karyotype is a fluorescencemicrograph of all the mitoticchromosomesin a cell (mitotic because the chromosomesare much morecondensed and visible than in interphase) ○ A chromosomeis 3-8 µm, while fluorescence microscopyhas a resolution of about 0.2 µm, so the approximateshapes and sizes of chromosomesare visible ○ Chromosomepairs all appear as different colors thanks to the use of special fluorescent dyes If chromosome2 is to be lime green, each lime green dye molecule is attached to a DNA sequence complementaryto a specific sequence found in nearly all chromosome2s in the species For this point, just understand that fluorescent dyes make the colors and the specificity is achieved by matching up sequences
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