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Biology Lecture :) > Exam 4 ;) > Flashcards

Exam 4 ;) Flashcards

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1
Q

Transcription

A

generation of RNA from DNA
requires a DNA template

Substrate: Nucleoside triphosphates (ATP, GTP, CTP, UTP)

Enzyme: RNA polymerase
No primer required
Prokaryotes – only one type
Eukaryotes – several types

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2
Q

transcription occurs in three steps

A

initiation
elongation
termination

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3
Q

Intiation

A

Requires a promoter (DNA sequence to which RNA polymerase binds)
Where RNA polymerase is to bind and which strand of DNA to transcribe
Transcription start site (where transcription begins)

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4
Q

Elongation

A

RNA polymerase: unwinds 13 base pairs at a time and reads template strand 3’ to 5’
Adds nucleotides at the 3’ end
Complementary base pairing
Ribonucleoside triphosphates (ATP, UTP, GTP, CTP) joined by phosphodiester bonds, releasing a pyrophosphate
DNA rewinds and RNA made as a single-strand
Proofreading?

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5
Q

Termination

A 
DNA sequence indicates end of the process 
Transcription ends:
RNA polymerase is released
RNA molecule is released 
May be influenced by many factors
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6
Q

Pre-mRNA

A

primary (first) mRNA transcript, that requires processing before it moves out of the nucleus

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7
Q

Exons

A

(expressed regions): region of pre-mRNA that remains in the mature mRNA

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8
Q

Introns

A

(interveining regions): those regions of the pre-mRNA that are not part of the mature mRNA

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9
Q

Pre-mRNA Processing

A

(making mature mRNA)
cutting introns out
splicing exons together

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10
Q

Prokaryotes transcription

A

Most of the genomic DNA is coding

mRNA is instantly made into mature mRNA

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11
Q

Eukaryotic Gene Processing

A

Prior to translation
RNA splicing
Addition of 5’ cap
Addition of poly A tail (3’)

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12
Q

RNA Splicing

A

removal of introns

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13
Q

snRNPs (small nuclear ribonucleoprotein particles

A

bind to consensus sequences of pre-mRNA
One binds near 5’ exon-intron boundary
One binds near 3’ exon-intron boundary

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14
Q

Proteins form spliceosome (RNA-protein complex)

A 
Cuts pre-mRNA at 5’exon-intron boundary
Intron forms loop structure
Cuts pre-mRNA at the 3’ exon-intron boundary
Releases introns (degraded in the nucleus)
Joins ends of exons together
Result = mature mRNA (exported from the nucleus for later translation)
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15
Q

Addition of a 5’ cap (G cap)

A

Modified molecule of GTP
Added to pre-mRNA as it is transcribed
Purpose:
helps mRNA bind to ribosome (preparation for translation)
Protects against digestion (ribonucleases – enzymes which break down RNA)

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16
Q

Addition of a poly A tail

A 
50-300 adenine nucleotides
Added to 3’ end of pre-mRNA
Purpose:
Helps with export of mRNA from the nucleus
Helps with stability of mRNA
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17
Q

Translation

A

Conversion of mRNA sequence into the amino acid sequence of a polypeptide (protein)
Change from the nucleic acid “language” into the amino acid “language

20 different amino acids 
    are encoded by the 
    nucleic acids
Side chains of amino acids
Unique functions
Increase characteristics
   of a polypeptide
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18
Q

Structure / Function of the tRNA

A 
Amino Acid Attachement Site:
Bind / carry particular amino acids (at 3’ end) 
Anticodon:
3 bases that bind mRNA
(noncovalent hydrogen bonds)
Interact with ribosomes:
3D structure of tRNA fits surface of ribosome
 (noncovalent hydrogen bonds)
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19
Q

Charging” of the tRNA

A

Aminoacyl-tRNA synthetases – family of 20 enzymes that required for attachment amino acids to tRNA

Each enzyme specific for one amino acid / tRNA group

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20
Q

Ribosome

A

site of translation

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21
Q

3 binding sites for tRNA

A

A (amino acid) site: region where new tRNA binds to mRNA via anticodon-codon bond
P (polypeptide) site: region where tRNA adds its amino acid to the polypeptide chain
E (exit) site: region where the tRNA (w/o amino acid) briefly resides before leaving the ribosome

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22
Q

Translation

A

initiation, elongation

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23
Q

Methionine

A

charged tRNA binds to AUG start codon

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24
Q

Steps of Translation

A

Codon recognition: anti-codon of tRNA binds to codon at A site
Peptide bond formation: (peptidyl transferase activity of the large subunit)
Elongation: free tRNA is moved to the E site and released; growing polypeptide chain moves to the P site
The process is repeated (until stop codon)

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25
Q

Translation: termination

A

Termination
Stop codon enters the A site (mRNA = UAA, UAG, and UGA)

Release factor binds to complex

Release factor disconnects polypeptide from tRNA in the P site (hydrolysis reaction)

mRNA and ribosomal subunits separate

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26
Q

Polyribosomes

A

purpose: increase rate of protein synthesis

groups of ribosomes on the same mRNA

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27
Q

Cellular destination of Proteins

A

Normally:
protein synthesis – begins with free floating ribosomes in cytoplasm…default end-location is cytosol
May contain signal sequence (short sequence of amino acids that indicates cellular location)

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28
Q

Some polypeptides are translated into the RER

A

Polypeptide with 5-10 hydrophobic amino acids at N-terminus — directed to RER
Polypeptide binds to receptor protein in RER membrane and translation continues
Signal sequence is removed
Translation continues till termination
Ribosome is released and protein folds inside of RER

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29
Q

Post-translational Modification of Proteins

A

Purpose: to influence function of the protein
Proteolysis – cutting of a polypeptide chain
Glycosylation – addition of carbohydrates to proteins (glycoproteins)
Phosphorylation – addition of phosphate groups (protein kinases

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30
Q

Proteolysis

A

cutting of a polypeptide chain

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31
Q

Glycosylation

A

addition of carbohydrates to proteins (glycoproteins)

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32
Q

Phosphorylation

A

addition of phosphate groups (protein kinases)

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33
Q

Constitutive Genes

A

expressed at all times

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34
Q

Inducible Genes

A

expressed only when needed

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35
Q

Receptor-Ligand Binding

A

Signal transduction

Gene activation vs. gene repression

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36
Q

Cell Cycle

A

Cyclins (bind CDKs and activate them, progression through the cell cycle)
Expression of cyclin genes at specific points during the cell cycle

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37
Q

Virus-infected Cells

A

“hijack” host gene expression machinery

Divert it to viral gene expression

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38
Q

When is gene expression regulated?

A

receptor-ligand binding
cell cycle
virus-infected cells

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39
Q

Where is gene expression regulated?

A

Transcriptional
Post-transcriptional
Translational
Post-translational

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40
Q

Gene expression is very precisely…

A

regulated

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41
Q

Transcriptional Regulation

A

Selective Gene Transcription
(Transcription Factors (TFs)
Repressors: (negative regulation) prevent transcription
Activators: (positive regulation) stimulate transcription

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42
Q

Repressors

A

negative regulation

prevent transcription

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43
Q

Activators

A

positive regulation

stimulate transcription

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44
Q

Viruses

A

– regulate gene expression to evade the host immune response
Acellular: depends on living cells to reproduce
Genome: dsDNA, ssDNA, dsRNA, ssRNA
Survival: hijacking host gene expression machinery

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45
Q

bacteriophage

A

bacterial virus
DNA or RNA genome
May have lysogenic phase

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46
Q

HIV

A

Causes AIDS (acquired immunodeficiency syndrome
Retrovirus
Enclosed in phospholipid membranes (from previous host)
Membrane proteins: help with fusion of viral PM and infection of host cell

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47
Q

Regulation of Translation

A

miRNA
Modification of the 5’cap
Translational repressor proteins

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48
Q

miRNA

A

inhibition of translation

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49
Q

Translational repressor proteins

A

Bind mRNAs and prevent attachment to ribosome

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50
Q

Proteosome

A

large protein complex that hydrolyzes target proteins

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51
Q

Ubiquitin

A

76 amino acid protein that targets other proteins for degradation

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52
Q

Alternative Splicing

A

generation of families of different proteins with different activities and functions from a single gene

53
Q

Inducible operon

A

turned off unless needed

54
Q

Repressible operon

A

turned on unless not needed

55
Q

Lac Operon

A

encodes lactose-metabolizing enzymes

Structure: 3 enzyme genes, promoter, operator
High rate of mRNA synthesis, when needed
No transcription, when not needed

56
Q

Prokaryotic Gene Regulation

A

Normally: only make the necessary proteins (conserve energy and resources)

57
Q

3 proteins that are critical for the uptake and metabolism

A

3 proteins that are critical for the uptake and metabolism:
β-galactoside permease – carrier protein in plasma membrane
β-galactosidase – enzyme that hydrolyzes lactose to glucose and galactose
Β-galactoside transacetylase – transfers acetyl groups from acetyl CoA to certain β-galactosides

58
Q

Lacatose is a

A

β-galactoside (a disaccharide with galactose and glucose)

59
Q

Lactose Metabolism in E. coli

A

Immediately begin making enzymes (3000/cell in 10min.):
β-galactoside permease
β-galactosidase
Β-galactoside transacetylase

60
Q

No Lactose Metabolism in E. coli

A

Low level (few molecules / cell):
β-galactoside permease
β-galactosidase
Β-galactoside transacetylase

61
Q

Operon

A

Cluster of genes with a single promoter that are transcribed together

62
Q

Two types of operons

A

Inducible

repressible

63
Q

Inducible Operon

A

turned off unless neede

64
Q

Repressible Operon

A

Turned on unless not needed

65
Q

Lac Operon (-)

A

(-) lactose
an inducible system
without lactose repressor is bound to operator sequence

66
Q

Lac Operon (+)

A

(+) lactose

presence of lactose

67
Q

trp operon

A

a repressible system

68
Q

Two types of chromatin

A

Euchromatin: loosely packed, undergoing transcription
Heterochromatin: tightly packed, not actively transcribed

69
Q

Euchromatin

A

loosely packed, undergoing transcription

70
Q

Heterochormatin

A

tightly packed, not actively transcribed

71
Q

Regulation of chromatin structure

A

Histone Acetylation: decrease binding to DNA

Histone Methylation: increase binding to DNA

72
Q

Histone Acetylation

A

decrease binding to DNA

73
Q

Histone Methylation

A

increase binding to DNA

74
Q

RNA polymerase binding stie

A

transcribes protein-coding genes

75
Q

General TF binding site

A

generatl transcription factors-bind to promoter site, allowing RNA polymerase 2 to next bind

76
Q

Gene-Specific TF

binding site

A

Bound by specific transcription factors
Activators – bind to enhancer sequences
Repressors – bind to silencer sequences

77
Q

Activators

A

bind to enhancer sequences

78
Q

Repressors

A

bind to silencer sequences

79
Q

Alternative splicing

A

Purpose: generation of families of different proteins with different activities and functions from a single gene

80
Q

microRNA (miRNA)

A

Purpose: degradation of mRNA
Small, noncoding RNAs
22 nucleotides in length
Dozens of mRNA targets
Made as a longer precursor, that is cleaved to make a double-stranded miRNA
First discovered in C. elegans (worm model used to study developmental biology)

81
Q

Regulation of Translation

A

miRNA – inhibition of translation
Modification of the 5’cap
mRNA capped with unmodified GTP = not translated
Translational repressor proteins
Bind mRNAs and prevent attachment to ribosome

82
Q

Post-translational Regulation:protein stability

A

Ubiquitin – 76 amino acid protein that targets other proteins for degradation
Proteosome – large protein complex that hydrolyzes target proteins

83
Q

Post-translational regulation

A

protein stability

84
Q

Ubiquitin

A

76 amino acid protein that targets other proteins for degradation

85
Q

Proteosome

A

large protein complex that hydrolyzes target proteins

86
Q

Prokaryotic Genomes

A

First genome to be sequenced (Haemophilus influenzae) by Craig Venter

87
Q

Eukaryotic genomes

A

Are larger than prokaryotic genomes

Have more regulatory sequences (proteins) than prokaryotes

Contain large amounts of noncoding DNA

88
Q

model organisms

A

reveal characteristics of eukaryotic genomes

used in the laboratory to determine characteristices that are broadly applicable

89
Q

Eukaryotic organisms contain

A

gene families

90
Q

Gene families

A

a set of similar genes derived from the same parent gene

91
Q

Pseudogenes

A

nonfunctional genes; arise from mutations that cause loss of function (may lack promoter or not splice properly)

92
Q

Eukaryotic Genomes contain

A

repetitive sequences

93
Q

Highly repetitive

A

Short (less than 100bp), repeated 1000’s of times in tandem
Genome: 10% (humans) to 50% (some species of fruit flies)
Often associated with heterochromatin
Short tandem repeats (STRs):
1-5bps, repeated up to 100X
Chromosomal location varies and is inherited

94
Q

moderately repetitive sequence

A

Repeated 10 – 1,000 times
Include genes for tRNAs and rRNAs (multiple copies)
Many are transposons…what are these?
40% of human genome

95
Q

Retrotransposons

A

make RNA copies of themselves, copied back into DNA, inserted in new genomic (class 1) locations

96
Q

DnA transposons

A

no RNA intermediate and no replication, excised from one location and inserted into another (Class 2)

97
Q

Transposon

A

sequences of DNA that move around within the genome

Might be inserted into a gene sequence — alternative mRNA / inactivated gene

Short sequences (1,000 to 2,000bp

98
Q

What are the types of sequences in eukaryotic genomes?

A

Single-copy Genes
Moderately Repetitive Sequences
Highly Repetitive Sequences

99
Q

The Human Genome Project

A

Public project completed in 2003
undertaken to determine the normal sequence of the human genome
determine mutations and relate them to phenotypes

100
Q

Overview of the Human Genome Project

A

3.2 billion bp in haploid genome = 24,000 protein-coding genes
Average gene: 27,000bp (1,000 to 2,400,000bp)
All genes have many introns
3.5% is functional, but noncoding — role in gene regulation (microRNA)
Over 50% is made of transposons and other repetitive sequences
Most (97%) is the same in all people

101
Q

Genomics

A

study of the genome

102
Q

Proteomics

A

study of the proteome

103
Q

Metabolomics

A

study of the metabolic intermediates and products an organism produces

104
Q

Human Genomics and Benefits for Medicine

A

Understanding of the genetic basis of disease

SNPs (single nucleotide polymorphisms) – single nucleotide variations; may vary between individuals or alleles

105
Q

DNA Microarray

A

may be used to determine which SNPs are associated with human disease (ie. breast cancer, diabetes, arthritis, obesity, and coronary heart disease)
Private companies: scan your genome for SNP alleles

106
Q

Pharmacogenomics

A

study of how an individual’s genome affects their response to drugs

Genetic variations affect how well an individual responds to a particular drug

Analysis used to predict whether or not a person will respond well to a drug

107
Q

Biotechnology

A

any technological application that uses biological systems, living organisms, or derivatives thereof to make or modify products or processes

108
Q

Recombinant DNA Technology

A

Single molecule, containing DNA sequences from two or more organisms

Four necessary tools:
Restriction enzymes (RE) – 
       cut DNA into fragments for 
       manipulation 
DNA ligase – joining DNA fragments together
Vector – carrier of recombinant DNA
Reporter Genes
109
Q

Recombinant DNA Technology Tools (4)

A 
Restriction enzymes (RE) – 
       cut DNA into fragments for 
       manipulation 
DNA ligase – joining DNA fragments together
Vector – carrier of recombinant DNA
Reporter Genes
110
Q

Restriction Enzymes

A

cuts dsDNA at specific sequences

111
Q

Making recombinant DNA from DNA fragments:_________

A

DNA Ligase

112
Q

Vectors

A

Carrier for Recombinat DNA

113
Q

Plasmids

A

Small circular DNA molecules
Autonomous replication within bacteria
Favorable because…
Small (2,000-6,000bp) – easy to manipulate
Contain one or more RE sites – easy insertion of DNA
Contain “resistance genes” – easy selection
Contain bacterial origin of replication – replication independent of host chromosome

114
Q

Viruses

A

accommodates many larger eukaryotice genes

infect cells naturally

115
Q

Expression Vectors

A

Include the appropriate sequences for transcription and translation

116
Q

Expression Vectors Prokaryotes

A

Promoter
Transcriptional termination signal
Sequence for ribosome binding

117
Q

Expression Vectors Eukaryotes

A 
Promoter (with transcription factor binding sequences)
Enhancers
Transcriptional termination signal 
Sequence for ribosome binding
Poly A addition sequence
118
Q

Reporter Genes

A

used to identify host cells with recombinant DNA

119
Q

Anitbiotic resistance genes

A

First: determine cells with plasmid
Second: determine cells with desired insert

120
Q

Green Fluorescence Protein (GFP)

A

Emits green light when exposed to UV light

Widely used

121
Q

Selectable Marker

A

a gene that can be used to identify cells that contain recombinant DNA

122
Q

Gel Electophoresis

A

separation of DNA fragments

The number of fragments:
How many times is the restriction site present in the DNA sample?
The size of fragments:
Use of DNA ladder (for size comparison)
The relative abundance of fragments:
Intensity of band
May use slice of gel with desired DNA for future experiments!

123
Q

DNA mutations can be made in the laboratory

A

Nature: mutations give cause and effect data
Problem?

Recombinant DNA mutations: more easily studied

Ex. Proteins associated with disease, hemoglobin, NLS (nuclear localization signals)

124
Q

Mutations may be studies in knockout

A

mice

125
Q

Homologous Recombination

A

exchange of segments between 2 DNA molecules, based on sequence similarity; similar sequences align and crossover

126
Q

Complementary RNA can be used to

A

prevent expression of specific genes

purpose: block translation of mRNA

microRNAs
Short, ssRNA that are complementary to target mRNA
Target mRNA degraded

127
Q

MicroRNAs

A

microRNAs
Short, ssRNA that are complementary to target mRNA
Target mRNA degraded

128
Q

Antisense RNA

A

Base pairs to mRNA
Partially dsRNA – inhibits translation
Used in anti-cancer therapy (reduced expression of genes associated with cancer)

129
Q

siRNA

A

Similar to microRNAs
Short (21-25nt) dsRNA molecules
Discovered in late 1990s

Biology Lecture :) (4 decks)

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