Notes for Second Midterm.docx

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Notes for Second Midterm
Folding is important since it reduces the loss of function, prevents diseases which are associated
with miss-folded proteins.
-helix – it is right handed helix stabilized by H bonds between C=O of peptide group n and N-H
of peptide group n+4.
3.6 AA are required for one complete turn. The R group points outwards.
Ala, Leu, Met, Glu (Normally found in  helix).
Pro, Gly, Ser, Thr (Normally not found in  helix and are referred to as helix breaker)
Hydrogen bond is between the O at carbonyl group to the H atom on the amid group on
the n+4 amino acid.
-sheet – adjacent chains held together by hydrogen bonds between
backbone. Stability depends on the number of hydrogen bonds. Can be
divided into three classes:
Parallel – chains run in the same direction (One side will have either
all N or C terminus, R group all point in same directions)  Bottom
Antiparallel – chains that run in opposite direction (one side will have
alternating patter of C and N and R groups will either be facing up or
down)  Top
Mixed – will contain combination of parallel and antiparallel.
5-10 AA per strand, and can have 2-15 strands. Results in formation of
either cylinder or barrel shape or flat planar sheets.
Note that the arrow head points towards the C-terminus
Motifs and domain are part of tertiary of structure.
Motif: recurring combination of secondary structure
in different region of the protein, ex (helix-turn-
helix). It is described in secondary structural terms.
The helix – loop – helix is a Calcium ion binding motif. It is
able to hold calcium ion because it has negatively charged
Asp, Asn, Thr.
Zinc-finger motif is designed to hold Zn ion. It holds the Zn
with help of 2 His pointing towards the Cys which holds the
Zn atom.
Domain: subunit of 3D structure which describes
how a protein will fold into an independent shape.
One polypeptide can have more than one domains, it
just depends on how many 3D structure it forms.
Domains contain secondary structure, thus it could
also have motifs as part of it.
Genotype: the genetic information that is encoded into the linear system of the nucleotides within
the DNA.
The primary structure of the DNA codes for the primary structure of the protein.
In Eukaryotes genetic info is segregated into different locations like the nucleus, mitochondria
and chloroplast. While in prokaryotes it’s present in nucleoid and plasmids which are integrated
with cytoplasm.
Gene is a region of DNA which controls a discrete hereditary trait of characteristic (genesis). It is
physical and functional unit of hereditary which carries information from one generation to next.
It is an entire DNA sequence necessary for the production of a functional protein or RNA
Molecule (Molecular view)
DNA codes for a message which is translated into an amino acid sequence of protein.
There is a switch which determines when and when not to transcribe the DNA.
Gene sintony is the order of which genes occur on chromosomes and they are used to control
cluster of genes along with individual genes.
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Genes are discrete unit, one gene will not
overlap with the other gene. The exception to
this is the viruses because they have small
genome.
The first amino acid in most of the protein
was methionine.
Gene NEEDS a start (promoter), stop
(terminator) and open reading frame (the part
which contains the information for creation
of the gene).
Not all the DNA in the organism codes for a gene. There are many other DNA sequences
between genes which doesn’t contain information for synthesis of proteins and RNA.
As the species get complicated the gene density decreases. Ex. Humans have gene density of
10% and 90% of the genes is non-coding gene, it was junk.
Aprox 500 gene is the bare minimum for any living organism.
Prokaryotic Gene:
It has a promoter – a region to which
RNA Pol binds. (non-coding region)
Exon – the region between the start and
stop which is transcribed into the RNA
and encodes most or all part of the
protein. UTRs: 5’ and 3’ UnTranslated
Regions, which are transcribed into RNA
however, they aren’t translated into the
protein.
Eukaryotic Gene:
It has exon, promoter, and UTRs.
However, it contains Introns which are
region of eukaryotic gene that doesn’t code for proteins but is transcribed to RNA and later
removed.
Gene expression is the process which involves transcription and translation of a discrete DNA
sequence.
There are two types of mRNA formed:
Monocistornic – contains single gene.
Polycistornic – contains more than one gene. (generally in bacteria)
DNA-dependent RNA synthesis is the
process by which the RNA is coded from
the DNA.
RNA is synthesized from DNA template
is complementary to the DNA template
The nucleotides are added from 5’  3’.
RNA synthesis is catalyzed by RNA
polymerase. It creates single stranded
DNA template and doesn’t require primer.
The products are called primary
transcript.
Prokaryotic cells = mRNA (can be
translated immediately)
Eukaryotic cells = heterogeneous
nuclear RNA (hnRNA) or pre-mRNA. The hnRNA undergoes post-transcriptional
process to become mRNA and then it translated into protein.
In transcription the (-) strand (non-coding = template) is used by the RNA Pol to synthesise sense
mRNA. However, the (+) strand (coding = non-template) isn’t transcribed since its carries
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information which the mRNA contains (same polarity, similar base pairs). If (+) strand is
transcribed, it results in antisense RNA.
Transcription start site is always labeled +1, and upstream (away from gene) from start site is
labeled -1, -2 and so on. While downstream (towards the gene) is +2, +3 and so on.
A DNA-RNA hybrid happens during transcription, however it is short lived. In this the RNA
forms complementary base pairs with DNA.
The process of RNA synthesis is self-energizing reaction since the energy required for this
process is provided by hydrolysis of the phosphate groups on each nucleotide base.
Eukaryotic hnRNA:
In the processing step, the introns are
removed through the process of splicing.
In this the sugar phosphate backbone is
broken into sections where the introns
are removed and the exons are ligated
together to form ssRNA molecule,
which contains the full gene. The
processing is done in the nucleus of the
cell.
Polycistronic message contains a
promoter which controls the
transcription of more than one gene. In
this ssRNA, there are more than on gene
present. The space between each gene is
called the ribosome binding site. This
site is used to regulate one or all of the protein regulation present on the ss mRNA.
The General Transcription in Prokaryotes:
The RNA Pol has the sigma factor attached to it which allows
the RNA Pol to attach to the start sequence. Once the RNA Pol
has been attached and the transcription has been started, the sigma
factor releases itself (after 10-15 nucleotide have been copied)
and binds to another RNA Pol. *Note that there are multiple types of
sigma factors, each associated with specific promoter region.
In prokaryotes the RNA Pol is a globular protein which consist of 5 subunits with a catalytic site.
Variable regulatory factor: sigma
RNA Pol holoenzyme = core + sigma factor.
The core enzyme consist of:
α-subunit x2 (36,000 Da x 2) – recognize the upstream element (-40 to -70) in DNA.
β-subunit (150,600 Da) – has the polymerase activity (catalyzes the synthesis of RNA)
which includes initiation and elongation.
β’-subunit (155,600 Da) – binds to the DNA and maintains the stability of the RNA Pol
ω-subunit (10,000 Da) – polymerase stability and helping the RNA Pol maintain its 3D
structure.
Sigma factor (between 7 to 60 different types of sigma factors) is the variable component
of RNA Pol holoenzyme. It tells that the specificity of the RNA Pol varies depending on
the sigma factor. Sigma 70 (70Da) – general house keeping sigma factor which is
responsible for transcribing RNA and respiration. Sigma 32 – heat stress response.
Transcription can be broken down into 3 stages:
Initiation: the binding of RNA Pol to a promoter sequence.
Elongation: addition of ribonucleotide to the RNA
Termination: dissociation of RNA Pol and release of the primary transcript from
template.
The control of transcription is important to production of protein within the cell. The is done by
controlling the life time of the RNA. Another the way is by increasing or decreasing the
transcription and translation.
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