40160754
| Questão | Responda |
| Prokaryotic RNA Polymerase | Core enzyme responsible for rapid elongation of RNA transcript Cannot initiate transcription without the sigma (σ) factor |
| Prokaryotic RNA Polymerase Structure | 5 Subunits: 2 alpha subunits, 1 beta subunit, 1 beta' (beta-prime) subunit, 1 omega subunit |
| RNA Polymerase 'Holoenzyme' | Prokaryotic RNA Polymerase with an additional subunit (sigma factor); responsible for transcription initiation |
| 3 Stages of Prokaryotic Transcription | Initiation Elongation Termination |
| Initiation STAGE I of PROKARYOTIC TRANSCRIPTION | RNA polymerase holoenzyme binds to a promoter after it scans along the chromosome, forming a closed complex; DNA is then unwound to be ss, forming an open complex |
| Typical Bacterial Promoter | Activates polymerization for transcription Contains two sequences; the "Pribnow box" at the -10 and -35 seq. (bound by sigma factor with phosphodiester bonds) |
| Elongation STAGE II of PROKARYOTIC TRANSCRIPTION | Sigma factor detaches from the prokaryotic RNA polymerase RNA chain is progressively elongated to synthesize an entire bubble |
| Termination STAGE III of PROKARYOTIC TRANSCRIPTION | Prokaryotic RNA polymerase falls of DNA Newly formed RNA transcript is released RNA transcription bubble is closed |
| Prokaryotic vs. Eukaryotic Transcription | Prokaryotes have no nucleus; thus transcription occurs freely in the cytoplasm -can even be translated simultaneously Eukaryotes must transcribe mRNA in the nucleus, modify it, then transport it to the cytoplasm for translation |
| Eukaryotic Modifications in Transcription to heterogenous nuclear RNA (hnRNA) *hnRNA refers to immature mRNA for eukaryotes | Splicing out introns Adding a 5' cap Adding a 3' Poly-A tail |
| Splicing Out Introns EUKARYOTIC MODIFICATIONS in TRANSCRIPTION | Mediated by the spliceosome (contains 100s of proteins and 5 snRNAs) Proteins bind snRNAs to form snRNP which bind to conserved areas of intron (5' GU, 3' AG, and A upstream 3') Intron is cut, attached to A, removed as a loop, while the two exons are joined |
| Addition of a 5' Cap EUKARYOTIC MODIFICATIONS in TRANSCRIPTION | Methylated guanine nucleotide is stuck on the 5' end, necessary for translation |
| Addition of a 3' Poly-A Tail EUKARYOTIC MODIFICATIONS in TRANSCRIPTION | A string of several hundred adenine nucleotides is added to the 3' end to prevent mRNA digestion by free exonucleases in the cell |
| Types of RNA Polymerases (Transcription) *Not to be confused with DNA polymerases in replication | Prokaryotic RNA Polymerase Complex (alpha, alpha, beta, beta', omega) In Eukaryotes; RNA Polymerase I, II, and III |
| RNA Polymerase I | Transcribes most rRNA |
| RNA Polymerase II | Transcribes: hnRNA, most snRNA, some miRNA |
| RNA Polymerase III | Transcribes: tRNA, Long ncNRA, siRNA, some miRNA, subset of rRNA |
| Translation | Synthesis of polypeptides according to the amino acid sequence, dictated by the sequence of codons in mRNA mRNA binds to ribosome, tRNAs deliver AA, bound together by ribosome; are released |
| Transfer RNA (tRNA) | Single RNA transcript produced by RNA polymerase III, responsible for delivery of amino acids to the ribosome for binding to create a polypeptide |
| Transfer RNA (tRNA) Structure | Stem-and-loop structure stabilized by hydrogen bonds; 5' end, D-loop, Anticodon Loop, T loop, 3' end (AA attachment site) |
| Wobble Hypothesis (tRNA) | Since there are 61 codon combinations, it would be expected that there are 61 tRNAs However, there's less than 45 -theorized to be attributed that while the first 2 bases of an anticodon match; the third may 'wobble' |
| Wobble Hypothesis Base Pairing | G - U I - U I - A I - C I is inosine, a modified base that helps with 'the wobble' |
| Amino Acid Activation / tRNA Loading | Peptide bond formation during protein synthesis is energetically unfavourable, thus requires HYDROLYSIS OF 2 HIGH-ENERGY PHOSPHATE BONDS to attach an AMINO ACID to its tRNA MOLECULE (i.e., SPECIFIC and ENERGETIC delivery) |
| Amino Acid Activation / tRNA Loading STEPS | a) AA attached to ATP, PPi (P2O7) leaves, forms aminoacyl-AMP (a-AMP) b) PPi is hydrolyzed to release energy to 2Pi c) tRNA loaded via destruction of energy a-AMP bond; produces AMP and a-tRNA |
| Aminoacyl-tRNA Synthetases | Enzymes specific to each amino acid Allows attachment of AA to AA acceptor site on the tRNA molecule (despite it always being CCA) based on 3D structure |
| Svedbergs | Sedimentation rate used for ribosomes; indicates how quickly something will sink in a gradient during centrifugation (units are not additive) |
| Prokaryotic Ribosomes (Svedbergs) | Prokaryotes have a 70S ribosome; large subunit of 50S, small subunit of 30S. |
| Eukaryotic Ribosomes (Svedbergs) | Eukaryotes have an 80S ribosome; large subunit of 60S, small subunit of 40S. |
| Large Unit of the Ribosome | Where ribozymic activity happens (i.e., where amino acids are all linked) |
| Binding Sites of the Complete Ribosome | A Site: Amino Acid delivery site (by tRNAs) P Site: Polypeptide chain's site of growth (where it is anchored to the tRNA) E site: Empty tRNA site prior to its ejection |
| Prokaryotic Translation | Synthesis of proteins using RNA transcript; occurs at the same time and in the same compartment as prokaryotic transcription (i.e., translation occurs as the RNA transcript is being made) |
| Sine-Dalgarno Sequence | Special site for binding of ribosomes (located at -10) |
| 3 Steps of Prokaryotic Translation | Initiation Elongation Termination |
| Prokaryotic Translation: INITIATION | small subunit (30S) binds to 2 initiation proteins (IF1, IF3); their complex binds the mRNA transcript; IF2 does the same, joins with fMet-TRNA (initiator tRNA ~a-TRNA), a modified Met is used as the first AA, forming an H-bond at the P site, not the A-site; large subunit (50S) joins; this whole process requires 1 GTP |
| Prokaryotic Translation: ELONGATION | second a-tRNA and form a H-bond to A site (requires GTP hydrolysis via EF-Tu), peptidyl transferase will help form peptide bonds between fMet and subsequent AA, translocation; each tRNA shifts over (requires a GTP); proteins are synthesized from N to C (akin to 5' to 3') |
| Prokaryotic Translation: TERMINATION | Once stop codon is reached, release factor enters the A site GTP is used to detach the ribosome |
| Eukaryotic Translation | Same steps as prokaryotic translation (initiation, elongation, termination), but: uses Met (not fMet), uses other sequence (not Shine-Dalgaro, but other 5' UTR ~Kozak), requires mRNA mod., not simultaneous, uses eIFs |
| Eukaryotic Initiation Factors (eIFs) | eIFs can be essential to initiating translation or helping regulate it can be reuglated via post-translation modifications (e.g., phosphorylation) |
| Cap Dependent Translation | Eukaryotic translation that starts at the 5' end of an mRNA because of the 5' cap requirement / recognition |
| Cap Independent Translation | Eukaryotic translation that may start in the middle of an mRNA molecule that have internal ribosome entry sites (IRES), typically involved with stress responses; its activation requires different proteins that normal initiation |
| Controlling Gene Expression (AT THE DNA LEVEL) | i) DNA Methylation and Chromatin Remodelling, ii) Gene Dosing, iii) Imprinting, iv) X-Chromosome Inactivation |
| Controlling Gene Expression (at the DNA Level): DNA METHYLATION + CHROMATIN REMODELLING | Methylation helps block transcriptional proteins from accessing the gene Methyl groups will also help attract other proteins that remodel the chromatin, winding it tighter, making it substantially harder to access |
| Controlling Gene Expression (at the DNA Level): GENE DOSING | Changing the number of copies of a gene would help control gene expression |
| Controlling Gene Expression (at the DNA Level): IMPRINTING | Only one allele of a gene is expressed (can change when passed to offspring) Gene can be silenced during imprinting using methylation, modifying histones, and binding long ncRNAs |
| Controlling Gene Expression (at the DNA Level): X CHROMOSOME INACTIVATION | In female mammals, only 1 X chromosome (referred to as Xa) is left active, while the other (Xi) is inactivated Inactivation is random; Xi will be very dense in heterochromatin and DNA methylation |
| Controlling Gene Expression (at the RNA Level w/ Prokaryotes) | Two key players: Lac Operon and Trp Operon |
| Operon | a functioning unit of DNA containing a cluster of genes under the control of a single promoter (seq. of nucleotides that activates RNA polymerization) |
| Lac Operon | inducible, catabolic enzyme - breaks down things and is only activated when object of interest (to be broken down is in excess) cleaves and transports lactose into cell given the absence of glucose |
| Lac Operon - Components | P region: promoter on DNA which RNA pol. binds to initiate transcription of Y, Z, A; O region: operator binding site of lac rep.; Z, Y genes: codes for enzymes that cleave / transport lactose, respectively A gene: codes for transacetylase |
| Lac Operon - Supplementary Genes | crp gene I gene |
| crp gene: lac operon - regulation (controls Z, Y, A) | codes for catabolite activator protein (CAP) (CAP helps couple the lac operon to glucose levels in the cell) |
| I gene: lac operon - regulation (controls Z, Y, A) | codes for the lac repressor protein (lac repressor protein prevents transcription of the lac operon) |
| Bacterial Cells preferred energy source? | Glucose |
| lac operon in the presence of glucose; activation state? | inactive |
| lac operon in the absence of glucose; activation state? | active |
| high glucose levels, means what for cAMP? (context of lac operon - normal conditions) | lower levels of cAMP; I gene's lac repressor protein is bound to O region of operon (in other words, the lac repressor protein is preventing lac operon activity ~transcription) |
| low glucose levels, means what for cAMP? (context of lac operon - high lactose levels) | higher levels of cAMP that bind to CAP; forms complex that binds to the operon's promoter and starts recruiting RNA pol. |
| Trp Operon | repressible, anabolic enzyme - breaks down things and is only activated when object of interest (to be broken down is in excess) (synthesizes tryptophan ~amino acid~ from chorismic acid) |
| Controlling Gene Expression (at the RNA Level w/ Eukaryotes) | Regulatory regions of protein coding genes, Enhancer sequences bound to activator pr, Gene repressor proteins (mod. chrom), Transcription factors (bound to DNA), RNA translocation, mRNA surveillance, and RNA interference |
| Controlling Gene Expression (at the RNA Level w/ Eukaryotes): Regulatory regions of protein coding genes | Upstream control elements, Core Promoter (contains elements for transcription complex / RNA pol. II), TATA box -where transcription complex assembly initiation occurs |
| Controlling Gene Expression (at the RNA Level w/ Eukaryotes): Enhancer Sequences | Enhancer sequences are bound to activator proteins |
| Controlling Gene Expression (at the RNA Level w/ Eukaryotes): Gene Repressor Proteins | Gene repressor proteins modify chromatin to regulate gene expression |
| Controlling Gene Expression (at the RNA Level w/ Eukaryotes): Transcription Factors | Transcription factors are bound to sections of DNA, impacting its transcription levels |
| Controlling Gene Expression (at the RNA Level w/ Eukaryotes): mRNA control | RNA translocation (controlling localization of mRNA), mRNA surveillance (elimination of low-quality transcripts), RNA interference (degrading mRNA via siRNA) |
| Post-Translational Modifications (context of Gene Expression) | Protein Folding, Covalent Modifications, Processing |
| Post-Translational Modifications: Protein Folding | Folding of newly synthesized proteins by chaperone proteins; chaperone proteins help proteins assume their native (correctly folded state), though mistakes may result in a non-native state |
| Post-Translational Modifications: Covalent Modifications | change of conformation, function, or target membrane; e.g., alkylated, formylated, sulphated, glycosylated, acetylation |
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