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Nucleic acids Flashcards

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1
Q

explain the significance of DNA

A

macromolecules needed for the storage and expression if genetic information
genes are made of nucleic acids and help us determine our characteristics
study faulty genes that can cause disease
DNA sequencing and genetic manipulation for research medicine and biotechnology

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2
Q

describe the structure of the subunits of DNA

A

made up from nucleotides
five carbon sugar - deoxyribose
nitrogenous base - ACTG (purines 2 or pyrimidines 1)
phosphate group

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3
Q

explain how double stranded DNA is formed

A

via phosphodiester bonds to form the sugar-phosphate backbone
hydrogen bonds holding the base pairs (pyrimidines and purines) together
double helical and antiparallel strands wrap around one another

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4
Q

describe the key features of double helical structure by most DNA molecules

A

right handed
diameter 2nm
height 3.4nm per turn
10 bp per turn
major and minor grooves

differences in DNA molecules are only in their nucleotide sequences

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5
Q

what is the difference between a nucleoside and nucleotide

A

nucleosides contain the sugar and the nitrogenous base whereas a nucleotide contains the sugar, nitrogenous base and the phosphate groups

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6
Q

how is a nucleoside formed?

A

when a base is linked to the 1’ carbon of a deoxyribose (or ribose) molecule

named deoxyadenosine, deoxycytidine, deoxyguanosine, deoxythymidine

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7
Q

describe some structural variants of DNA

A

B-DNA - common form
A-DNA - double stranded RNA, 11bp per turn, right handed slant and forms when DNA is dehydrated
Z-DNA - 12bp per turn, left handed, zig-zag backbone, may form if DNA contains long runs of alternating G and C

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8
Q

describe factors that influence denaturation of DNA

A

70-110 degrees, or exosed to alkaline conditions
strands will separate
if allowed to cool slowly, it can reanneal

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9
Q

explain how RNA and DNA differ

A

RNA has ribose instead of deoxyribose (making RNA more reactive and less stable)
RNA contains adenine, cytosine, guanine and uracil
RNA is usually single stranded
RNA resembles A-DNA not B-DNA
has more functions including, carrying info (mRNA), transporter (tRNA), structurally or catalytic (rRNA) and acts as a regulator of gene expression (micro RNAs)

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10
Q

gene

A

the entire stretch of DNA necessary for the production of a particular functional product, which may be a protein or an RNA molecule

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11
Q

what is underwinding and overwinding?

A

underwinding - negative supercoils
overwinding - positive supercoils

DNA becomes supercoiled when it is unwound, and is usually in vivo negatively supercoiled

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12
Q

how can denaturing be monitored?

A

measuring absorbance under UV light at a wavelength of 260nm
UV rises as DNA denatures (hyperchromic shift)

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13
Q

what determines melting temperature?

A

the temperature needed to denature 50% of the DNA molecules in a sample.
DNA with high content of GC base-pairs due to more H-bonds being present
presence if cations, reduce repulsions between negatively charged phosphate groups on the two strands

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14
Q

genome

A

the genetic material contained in an organism, cell, virus or organelle

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15
Q

chromosome

A

a single long molecule of DNA that includes numerous genes. The DNA of a chromosomes is usually associated with proteins. Eukaryotic chromosomes are visible during cell division

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16
Q

chromatin

A

the DNA-protein complex present in the nuclei of eukaryotic cells during interphase

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17
Q

bacterial genomes

A

only one chromosome and is circular
compressed to form nucleoid
may contain plasmids

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18
Q

plasmids

A

1kb-400kb
may contain 1 to 200 copies of plasmid in cell
non-essential genes
readily passes from cell to cell
original replication
exploited in genetic manipulation

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19
Q

outline the composition of the genomes of eukaryotes

A

mitochondrial genome - 16500bp in length, 22 encoding tRNA, 2 encoding rRNA, 13 encoding mRNA

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20
Q

explain how DNA is packaged into chromosomes in eukaryotes

A

1/3 DNA
1/3 histone proteins
1/3 non-histone proteins
nucleosomes - think beads on a string, the string is DNA, the beads are nucleosomes, 146bp and wrapped twice around the core containing:
2 molecules histone H2A
2 molecules histone H2B
2 molecules histone H3
2 molecules histone H4

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21
Q

is DNA replication conservative or semi-conservative?

A

semi conservative

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22
Q

packing ratio equation

A

packing ration = length of DNA/ length of structure DNA packed into

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23
Q

what are the types of histone proteins?

A

H1
H2A
H2B
H3
H4
20% of amino acids are arginine or lysine

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24
Q

function of H1

A

binds to DNA outside the core particle and seals DNA to nucleosome

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25
how are nucleosomes packed?
packed into the nucleus of interphase cells in disordered chains with varying density of nucleosomes
26
why is the mitotic chromosome useful?
it has a scaffold of non-histone proteins that anchor the long loops of nucleosomes the scaffold contains topoisomerase 2 and condensin (ring shaped protein that anchors the ends of a loop of DNA)
27
topoisomerase 2
an enzyme that can remove or add supercoils within DNA
28
what does histone acylation do?
makes DNA less tightly associated with nucleosome and more accessible for transcription so that the gene may be switched on
29
describe the mechanism of DNA replication in E.coli in terms of replication forks
the origin of replication is rich in A and T, there are two replication forks bidirectional replication NOTE: eukaryote chromosomes have multiple origins of replication
30
what are the names of the DNA polymerases that E.coli uses? There are 5 but name 2
DNA polymerase I DNA polymerase III
31
what does DNA polymerase need in order to synthesise DNA?
all four deoxynucleoside triphosphates a template a primer
32
DNA polymerase I: what does 5' 3' exonuclease allow alongside DNA polymerase? what does 3'5' exonuclease allow?
nick translation (only for DNA polymerase I) DNA polymerase I and III to remove incorrect nucleotides from newly made DNA for proof reading
33
describe DNA replication in E.coli
partial unwinding using helicase (positive supercoils are removed by DNA gyrase and type II topoisomerase) synthesis of primers by primase (RNA primers) DNA synthesis by DNA polymerase III, the leading strand is synthesised continuously and the lagging strand is synthesised in Okazaki fragments removal of RNA primers by nick translation by DNA polymerase I sealing the gaps between Okazaki fragments by DNA ligase
34
semi-discontinuous
lagging strands synthesised as Okazaki fragments
35
explain how transcription occurs in E.coli
initiation - RNA polymerase binds to the promoter (identified using -35 hexamer and pribnow box - consensus sequences) , DNA unwinds and RNA synthesis begins elongation - RNA polymerase moves along the DNA molecule synthesising an RNA copy (15-17bp at a time) termination - RNA polymerase dissociates from DNA releasing the new RNA molecule
36
what forms of RNA polymerase in E.coli are there?
holoenzyme - hexamer with the subunit structure alpha2betabeta'omegasigma, which carries out initiation but not elongation this binds to the promoter with the sigma subunit dissociating from the core enzyme core enzyme - pentamer with the subunit structure alpha2betabeta'omega which carries out elongation and initiation after termination the core enzyme dissociates from DNA and binds a sigma subunit to reform the holoenzyme
37
how does transcription in eukaryotes differ to that in prokaryotes?
eukaryotes have separate RNA polymerases for mRNA, rRNA, and tRNA eukaryotes make a primary transcript (pre-mRNA) that is processed in the nucleus to form mRNA eukaryotic promoters differ from those in bacteria eukaryotic RNA polymerases cannot directly recognise the promoter
38
name the three types of RNA polymerases in eukaryotes
RNA polymerase I - synthesis rRNA RNA polymerase II - synthesis mRNA RNA polymerase III - synthesis tRNA
39
describe RNA processing in eukaryotes
the primary transcript is processed (pre-mRNA) by: capping - addition of methylated G to 5' end to stabilise the mRNA molecule used for translation cleavage and polyadenylation - 3' end of the primary transcript is cleaved and a 'poly A tail' 250A residues is added to stabilise the mRNA molecule RNA splicing - exons are spliced together to generate mRNA
40
transcription in eukaryotes by RNA polymerase II
promoter recognised and require accessory proteins including transcription factors. the core promoter interacts with the general transcription factors to form the pre-initiation complex for the initiation of transcription core promoter including TATA box or more proximal promoter elements (upstream promoter elements) may also require enhancers for efficient transcription
41
describe the nature and operation of the genetic code
three nucleotides create a codon which an AA can be assigned to genetic code tells us which codon represents which AA features: non-overlapping degenerate universal
42
describe the structure of transfer RNA (tRNA)
73-90 nucleotides long unusual bases arising from post-translational modifications anticodon present upaired sequence CCA at 3' extensive internal base pairing cloverleaf structure with 3 or 4 loops tertiary structure presents an L
43
explain how transfer RNAs become linked to an AA
called aminoacyl tRNAs (charged tRNAs) amino acid becomes activated can have base wobble where 5' end of anticodon can pair with more than one base at the 3' end of the codon
44
what are the enzymes that link tRNAs to amino acids called?
aminoacyl-tRNA synthetases tRNA are joined to their amino acids in a 2-step ATP driven reaction catalysed by aminoacyl tRNA synthetases, an aminoacyl-AMP forms as an intermediate in the reaction
45
describe the structure of a bacterial ribosome
large subunit 50S and small subunit 30S and the whole ribosome is 70S the small subunit binds to the mRNA being translated, the large subunit catalyses peptide bond formation has an E, P and A site for the tRNA binding sites E = exits site P = peptidyl site A = aminoacyl site
46
describe translation
initiation: mRNA is read from 5' to 3' protein is synthesised from NH3+ to COO- in prokaryotes the first AA is N-formyl-methionine instead of methionine elongation: addition of amino acids to growing peptide termination: dissociation of the completed protein from the ribosome
47
compare and contrast translation in prokaryotic and eukaryotic organisms
eukaryotic ribosomes are larger than prokaryotic ribosomes initiation is different by recognition of ribosomal binding site (prokaryotes) or cap (eukaryotes) elongation is similar eukaryotic are prokaryotic ribosomes are sensitive to different antibiotics and inhibitors e.g: erythromycin/ clarithromyosin for prokaryotic large subunit inhibits the peptidyl transferase and translocation reaction
48
describe the structure of the human ribosome
overall size 80S large subunit is 60S and the small subunit is 40S note that mitochondria have their own ribosome 55S
49
what is the elongation cycle (repeated cycle)
binding of aminoacyl-tRNA needing GTP the peptidyl transferase reaction (peptide bond synthesis) translocation (movement of the ribosome one codon along the mRNA needing GTP) cycle repeats until the termination codon is reached
50
describe translation initiation in E.coli
small ribosome subunit binds to ribosome binding site on the mRNA as a result of base pairing between it and the 16S rRNA f-met-tRNAmet binds to the initiation codon 50S subunit binds, generating the complete ribosome
51
describe initiation in eukaryotes
first AA is methionine not N-formyl-methionine 40S subunit and met-tRNAmet combine prior to binding to mRNA no ribosome binding site on mRNA, recognition of the cap at the 5' end of the mRNA 40S subunit and met-tRNAmet migrate along the mRNA until the first AUG is reached, the 60S subunit then binds generating an intact ribosome with the first tRNA in the P site.
52
what is peptide bond formation catalysed by?
23S rRNA
53
ricin
antibiotic/ inhibitor eukaryotic 60S subunit target catalytic, removes a base from rRNA lethal dose is approx. 10-4g
54
outline the importance of regulation of gene expression
some genes may need to be switched on or off housekeeping genes on tissues specific genes in eukaryotes inducible genes in eukaryotes (hormonal activated genes) inducible genes in prokaryotes (expressed only when product is required so turned off so energy isn't wasted)
55
describe how the genes required for lactose metabolism are regulated in E.coli
when lactose is present alone: cAMP is abundant due to low glucose levels CAP-cAMP complex to CAP site stimulates binding of RNA polymerase to promoter the lac repressor is unable to bind to operator when complexed to allolactose (lactose to allolactose) so the strong transcription of lac Z, lac Y and lac A to produce beta galactosidase, permease and transacetylase in high concentrations
56
what are housekeeping genes?
encode products that are needed constantly so they are expressed at times and in all tissues
57
what can gene expression be controlled by?
regulating transcription (most common) regulating RNA processing (splicing of exons) regulating translation
58
operon
a cluster of several genes transcribed into a single mRNA common in prokaryotes but not eukaryotes
59
describe how the genes required for lactose metabolism when glucose is present are regulated in E.coli
cAMP conc is low due to high glucose concs RNA polymerase can transcribe genes but levels of transcription low because promoter is weak lac pressor unable to bind to operator when complexed with allolactose so low amounts of product formed from translation due to weak transcription of lac Z, lc Y and lac A
60
how do you determine strong and weak promoters?
the more a promoters actual gene differs from the consensus sequences, the weaker the promoter the lac promoter is quite weak due to it differing from the ideal promoter in three places
61
describe how the genes required for lactose metabolism when lactose is absent are regulated in E.coli
RNA polymerase cannot bind to promoter, no transcription occurs operon is repressed
62
what is the lac repressor protein encoded by?
Lac i gene
63
when lactose is present, some of it is converted into what isomer?
allolactose
64
what is catabolite repression?
when the lac operon genes are transcriibes weakly because both lactose and glucose are present the removal of catabolite repression requires: catabolite activator protein (CAP) and cyclic AMP
65
describe the control of gene expression in eukaryotes
core promoters and proximal promoter elements (PPEs) and also influenced by distal regulatory sequences called enhancers expression of eukaryotic genes requires the binding of PPEs and transcription factors

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