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BCHM 4360 EXAM 3 ACTUAL EXAM 2025 | ALL
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What is RNase E and what does it do? ---------CORRECT ANSWER---------- -------Endonuclease that is responsible for degradation of bacterial mRNAs, part of the degradosome complex How does eukaryotic RNA degradation work? ---------CORRECT ANSWER- ----------------3' poly(A) tails hinder rather than aid degradation in eukaryotes
- the first step in degradation is often tail shortening by a deadenylase.
- After this, decapping enzymes remove the 5' cap and 5' to 3' exonucleases degrade the RNA, or the exosome catalyzes exonuclease activity in the 3' to 5' direction Describe what happens to transferrin receptors when iron is low and when it is high. ---------CORRECT ANSWER-----------------High - mRNAs encoding the receptor are degraded as there is no need to import more iron Low - receptor mRNA is protected from degradation by Iron Regulatory Proteins (IRP1/IRP2) that bind to stem-loop Iron Response Elements (IRE) Foreign nucleic acids are removed by RNA interference (RNAi) in __________ and CRISPR interference in __________. ---------CORRECT ANSWER-----------------eukaryotes, bacteria What happens to defective endogenous RNAs? ---------CORRECT ANSWER------------------In eukaryotes, ribosomes on defective RNAs are marked by interaction with proteins such as the EJC, which recruit RNAses to degrade the RNA
- In bacteria, stalled ribosomes are recognized by a complex containing tmRNA, which allows the ribosome to finish translation using the tmRNA as a template (also recruits RNases) What is the RNA-recognition motif (RRM)? ---------CORRECT ANSWER----- ------------Also known as RNA-binding domain (RBD) or ribonucleoprotein (RNP) domain, this is where the RNA binds proteins
- Has alpha helices and four beta sheets in a sandwich, very adaptable and can bind RNAs that have different structures What are the differences and similarities between miR:miR* and siR:siR* molecules? ---------CORRECT ANSWER-----------------Both have 3' OH and 5' monophosphates and both can undergo post-transcriptional modifications such as methylation siR:siR* molecules are usually fully complementary, but miR:miR* molecules are usually not What are the three main classes of Argonaute proteins? ---------CORRECT ANSWER-----------------1. Argonaute proteins involved in miRNA and siRNA mechanisms in animals, plants, and fungi
- Piwi subclass involved in piRNA mechanisms
- WAGO proteins found in C. elegans What are the four domains of Argonaute proteins and what do they do? ----- ----CORRECT ANSWER-----------------1. PAZ binds the 3' end of bound RNA
- Mid interacts with the 5' end of the RNA
- PIWI is related to RNAse H and interacts with the whole RNA
- PAZ, Mid, and PIWI together orient the bound guide RNA to facilitate its scanning of cellular molecules
Protein folding can start as soon as the ___ terminus emerges from the ribosome, where the peptide encounters __________. ---------CORRECT ANSWER-----------------N, chaperones How do chaperones work? ---------CORRECT ANSWER----------------- interacting with hydrophobic regions on the emerging peptide, stopping them from making incorrect associations with other hydrophobic regions How do chaperones, like Hsp70 or DNAK work? ---------CORRECT ANSWER-----------------The nucleotide binding domain (NBD) hydrolyzes ATP and the substrate binding domain clamps a hydrophobic region of the polypeptide chain
- Co-chaperones like DNAJ and GrpE help bind the peptide or regenerate ATP and assist proper protein folding What are two cellular catalysts that aid disulfide bond formation? --------- CORRECT ANSWER-----------------1. Sulfhydryl oxidation makes new disulfide bonds
- Isomerization swaps disulfide bonds What is the difference between bacteria and eukaryotes with regards to oxidizing disulfide bonds? ---------CORRECT ANSWER----------------- Bacteria use DsbA sulfhydryl as the oxidase and DsbC as the isomerase, whereas eukaryotes use Protein Disulfide Isomerases to perform both functions Proteins can have multiple _______ motifs that fine-tune _____________. - --------CORRECT ANSWER-----------------sorting, localization
What are the three requirements for sorting proteins into cellular compartments? ---------CORRECT ANSWER-----------------1. Sorting motif recognition
- Transport
- Movement into or across a membrane The bacterial secretion system is very similar to ____ targeting in eukaryotes, relying on ______s to recognize and bind hydrophobic signals before handing proteins over to a translocase. ---------CORRECT ANSWER-----------------ER, SRPs True or False: Transport in and out of the nucleus is highly regulated and travel is via nuclear pore complexes (NPCs) ---------CORRECT ANSWER--- --------------True What are inteins? ---------CORRECT ANSWER-----------------Parts of proteins that catalyze their own removal and ligate the flanking pieces
- They often act as a mobile genetic element, moving its own DNA sequence around the genome What are the 4 steps to intein removal? ---------CORRECT ANSWER--------- --------1. A conserved residue (C, S, T) attacks the carboxyl of the previous peptide bond
- The same carboxyl is then attacked by a similar residue in the C-terminal exein
- The amino group of the last intein residue attacks its own carboxyl, separating the intein
- The two joined exeins undergo further attack of amino groups on carboxyl to make a peptide bond
5' trimming of tRNAs is done by which endonuclease? ---------CORRECT ANSWER-----------------RNase P True or False: Both the bacterial and eukaryotic RNA component of RNase P can cut RNA alone ---------CORRECT ANSWER-----------------False: Only bacterial can cut alone True or False: Some rRNA introns can catalyze their own removal and are subsequently called self-splicing. ---------CORRECT ANSWER----------------- True What is the CCA sequence? ---------CORRECT ANSWER-----------------It is the attachment site on the tRNA for the amino acid What are the two most common rRNA modifications? ---------CORRECT ANSWER-----------------1. ribose 2'-O-methylation
- pseudouridylation True or False: Post-transcriptional chemical modifications are limited to small additions such as methylation ---------CORRECT ANSWER-------------- ---False: Modifications can be small, like methylation, but can also be large, like the addition of a threonine What are snoRNAs and what is their general purpose? ---------CORRECT ANSWER-----------------small nucleolar RNAs typically made from introns of precursor mRNAs and they are typically used as guide RNAs
Describe how snoRNAs catalyze methylation of nucleotides --------- CORRECT ANSWER-----------------The snoRNA base-pairs with specific regions of target RNAs and directs enzymes to these positions snoRNAs that direct ribose methylation are _______ snoRNAs. snoRNAs that direct pseudouridylation are ______ snoRNAs. ---------CORRECT ANSWER-----------------box C/D, H/ACA What is the primary purpose of the end modifications of eukaryotic RNAs following transcription? ---------CORRECT ANSWER-----------------protection of mRNAs from nuclease degradation and assisting with protein interactions What are the 5' ends of mRNAs capped with? ---------CORRECT ANSWER- ---------------- 7 - methylguanine nucleotide via a 5'-5' triphosphate linkage What is the 5' cap needed for? ---------CORRECT ANSWER----------------- efficient elongation and termination of the transcript, for mRNA processing and export from the nucleus, and for directing translation What are the three steps for the addition of the 5' cap? ---------CORRECT ANSWER-----------------1. An RNA 5' triphosphate catalyzes removal of a phosphate from the 5' end
- A guanyl transferase attaches a guanosine monophosphate (GMP) to the end in a 5'-5' triphosphate linkage
- The guanine is methylated by a guanine- 7 - methyltransferase
True or False: Transcription, processing, and translation all occur in the nucleus. ---------CORRECT ANSWER-----------------False: Transcription and processing occur in the nucleus, but translation occurs in the cytoplasm so mature mRNAs must be exported from the nucleus What determines when the RNA-protein complex can be released from the transcription complex for export to the cytoplasm? ---------CORRECT ANSWER-----------------Protein factors needed for transport are loaded onto the mRNA during transcription, but polyadenylation is what is mainly needed Most introns do not contain genes and are excised and degraded. What are the two exceptions? ---------CORRECT ANSWER-----------------snoRNAs and certain miRNAs What general reaction do all mechanisms of intron removal use? --------- CORRECT ANSWER-----------------transesterification reactions True or False: Introns are far more prevalent in eukaryotes than in bacteria. ---------CORRECT ANSWER-----------------True What is the main benefit of having introns? ---------CORRECT ANSWER---- -------------They allow for exon shuffling, where exons are exchanged and reordered via recombination, allowing evolution of different genes. In addition, differential removal of introns gives different transcripts from the same gene
What is the mechanism of intron removal and how does it affect the overall energy? ---------CORRECT ANSWER-----------------Two step reaction, both involving transesterification reactions. First, the intron is detached from exon 1 by breaking the phosphodiester bond. Next, exon 1 reacts with exon 2 to replace the phosphodiester bond. The energy is the same in both bonds, so the reaction does not require ATP and is easily reversible. What are some characteristics of Group I introns? ---------CORRECT ANSWER-----------------Found in bacteria, viruses, microeukaryotes, and plants. They are ~120-450 nucleotides long Many (but not all) can excise themselves from the primary transcript. What is the mechanism of intron removal for Group I introns? --------- CORRECT ANSWER-----------------In the first transesterification, a free guanosine is the attacking species that detaches the 5' end of the intron from exon 1. In the second reaction, the released end of exon 1 attacks the intron-exon 2 junction, joining the two exons together What are some characteristics of Group II introns? ---------CORRECT ANSWER-----------------Found in bacteria and in organellar genes of plants and fungi ~400-1000 nucleotides long Can also act as mobile genetic elements by reverse splicing, and they often have open reading frames within the introns that assist splicing What is the mechanism for intron removal for Group II introns? --------- CORRECT ANSWER-----------------The 2' OH of a specific A within the intron attacks the exon1-intron junction.
What are the 5 steps of how the spliceosome works? ---------CORRECT ANSWER-----------------1. The 5' splice site is recognized by U1 and other sequences are bound by non-snRNPs. 2 - 3. The rest of the splicing apparatus binds, sometimes displacing other factors, including the branch-point-binding protein (BPP).
- The pre-mRNA rearranges, the first transesterification reaction occurs and the lariat forms.
- Other rearrangements then bring the two exons together for the second transesterification. What is the EJC? ---------CORRECT ANSWER-----------------exon junction complex, left at splice junctions after splicing and marks the transcript as processed and interacts with export and translation proteins What is the difference between cis and trans splicing? ---------CORRECT ANSWER-----------------Cis splicing is where exons in the same molecule are joined together. Trans splicing is unusual, and joins exons from different RNA molecules. PRP8, ATPases, and SR proteins are all proteins involved in splicing. What do they do? ---------CORRECT ANSWER------------------PRP8, found near the active site of the assembled spliceosome, thought to be important in catalysis.
- ATPases, which probably help with structural rearrangements and promote lariat and mRNA release after processing.
- SR proteins (named after serine/arginine repeats) are thought to help recruit spliceosome components in the splice sites What is alternative splicing? ---------CORRECT ANSWER----------------- different combinations of exons can be used, yielding more than one mature mRNA
Most exons are ____________ (must be included), but some are __________ exons (can be included or not) ---------CORRECT ANSWER--- --------------constitutive, regulated What are cryptic splice sites? ---------CORRECT ANSWER----------------- False splice sites that occur as a result of the simplicity of the sequences defining intron-exon junctions What are the two methods for how the spliceosome recognizes only true splice sites and not cryptic splice sites and how do they work? --------- CORRECT ANSWER-----------------1. Exon definition (mammals)
- The 5' and 3' ends of an exon are brought together by interactions between the U1 and U2 complex. Mutations at a 5' splice site would result in exon exclusion. The exon is typically marked with SR proteins and the introns bound with hnRNPs, which masks cryptic splice sites.
- Intron definition
- Introns are defined by interactions between 5' and 3' bound factors. Mutations at a 5' splice site would reslut in intron inclusion. Describe the similarities between the exon definition and intron definition models? ---------CORRECT ANSWER------------------In both models, the final splicing complex must be one in which the 5' and 3' complexes interact with one another across the intron
- In both models, the 5' and 3' splice sites are marked as they are transcribed, facilitated by RNA Polymerase and splicing components RNA sequence elements that positively affect splicing are called splicing _________. _________ sequences have the opposite effect. Both can
True or False: Normal RNAs are degraded the same way as foreign and defective RNAs. ---------CORRECT ANSWER-----------------False: They are degraded differently. RNA half-lives range from <1 minute to an hour in E. coli and ~20min to
24 hours in vertebrates. Why? ---------CORRECT ANSWER----------------- Some RNAs, like rRNAs, are needed a lot and are fairly stable. Others, like some mRNAs, are only required for short periods of time and so are rapidly degraded. What are the three main factors that contribute to RNA stability? --------- CORRECT ANSWER-----------------1. 5' cap in eukaryotic mRNA (also bacteria with a 5' triphosphate are more stable than those with a monophosphate)
- Poly(A) tail - increases stability in eukaryotes but decreases stability in bacteria
- Step loop structures What are AU-rich elements, where are they found, what do they do, and give an example? ---------CORRECT ANSWER-----------------AU-rich elements (AREs) are found in the 3' UTRs of some mRNAs, they are important for mRNA stability, they direct poly(A) removal and increase mRNA stability. c-fos in vertebrates encodes an ARE-containing short-lived transcription factor that promotes cellular growth, also some tumor-causing viruses express v-fos, which does not have the ARE (it is not degraded properly and leads to excessive cell growth)
Both bacteria and eukaryotes use what kind of system for RNA degradation? ---------CORRECT ANSWER-----------------endonuclease activity What are KH and PAZ? ---------CORRECT ANSWER-----------------ssRNA binding motifs
- KH domains have an RNA-binding groove formed by two alpha helices and one beta strand
- PAZ domains have a pocket formed from beta strands and an alpha helix (this pocket interacts with the single stranded overhangs of the siRNA
- PAZ domains are found in many RNAi proteins like Dicer What are unique about RNase III family enzymes? ---------CORRECT ANSWER-----------------Have dsRNA-binding domains
- Alpha helix and two loop regions that interact with three grooves on the dsRNA True or False: RNA-binding proteins often have more than one binding motif. ---------CORRECT ANSWER-----------------True What are three examples of RNA-binding motifs? ---------CORRECT ANSWER-----------------1. The eukaryotic PUF protein family, including Pumilio 1, has repeats of a two-helix motif with conserved amino acids, each interacting with a single nucleotide
- Some RNA-binding proteins need to have small regions rich in lysine and arginine
- Some proteins with the zinc finger DNA-binding domains also bind to RNA Amino acids are attached to tRNAs by ____________-____ ___________. ---------CORRECT ANSWER-----------------aminoacyl-tRNA synthetases
True or False: The first two positions on the mRNA are read by strict Watson-Crick base-pairing with positions 2 and 3 of the anticodon --------- CORRECT ANSWER-----------------True What is wobble pairing and what does it mean with regards to tRNA? --------
- CORRECT ANSWER-----------------The third position of the codon allows some pairing deviations, means that non-Watson-Crick interactions can occur and thus that each codon does not need its own tRNA (40 tRNAs for the 61 codons) Why do similar amino acids have similar codons? ---------CORRECT ANSWER-----------------Mutations that change the amino acid that is encoded will result in an amino acid with similar characteristics which generally won't affect the resulting protein too much _______________ attaches amino acids to tRNAs. ---------CORRECT ANSWER-----------------aminoacylation True or False: aminoacylation does not require ATP. ---------CORRECT ANSWER-----------------False What is the two-step mechanism for aminoacylation? ---------CORRECT ANSWER-----------------1. The amino acid is activated by attachment of AMP, releasing pyrophosphate and providing energy.
- Enzyme then transfers the amino acid to the 2' or 3' OH of the ribose of the terminal adenosine on the tRNA 3' CCA tail
The correct amino acid for a tRNA is referred to as _______. --------- CORRECT ANSWER-----------------cognate Loading of amino acids onto tRNA is very _______ with fewer than one error per _____ aminoacylation events. ---------CORRECT ANSWER--------- --------accurate, 100000 How do aminoacyl-tRNA synthetases recognize tRNAs? ---------CORRECT ANSWER-----------------sequence and structural features called identity elements How does the editing site prevent non-cognate amino acids from binding? - --------CORRECT ANSWER-----------------It can accomodate the pre-transfer amino acid, in which the activated amino acid is hydrolyzed if it is incorrect, or post-transfer, in which the amino acid is cleaved from the tRNA. What is the main difference between Class I and Class II aminoacyl tRNA synthetases? ---------CORRECT ANSWER-----------------Class I usually recognizes the minor groove of the acceptor stem, Class II the major groove
- Class I attach amino acids to the 2' OH of the terminal ribose, class II to the 3' OH Explain how some bacteria can operate with fewer than 20 synthetases (use glutamine and asparagine) ---------CORRECT ANSWER----------------- Aspartate and glutamate synthetases have dual specificity and a transamidase reaction changes the side chain from an acid to an amide
