BSC 310 Exam 2

Nucleotide structure

A nitrogenous base, connected to 1’ of a sugar (deoxyribose), and connected at the 5’ to a phosphate group

DNA structure

A complex structure of multiple nucleotides, with a antiparallel strand connected by hydrogen bonds to make the double strand.

RNA structure

A single strand of nucleotides, with Uracil instead of Thymine, and a ribose sugar

Negative Supercoiling

when the DNA strand is twisted in the opposite direction the double helix is twisted in (gets looser)

Positive Supercoiling

when the DNA strand gets twisted in the same direction of the double helix, and gets together. Used in prokaryotes to compact DNA

Central Dogma

the steps in biological information flow. DNA transcribed to RNA and then translated into protein

Plasmids

are free DNA in prokaryotes. can be circular or linear

Transposable elements

DNA sequences that can insert into other DNA and change its position about the genome

Steps of DNA replication (simple)

1) DNA Helicase unwinds strands

2) DNA Primase adds RNA priming sequence

3) DNA Polymerase III adds nucleotides in 5’ to 3’ direction

4) DNA Polymerase I removes primers, and adds DNA

5) DNA Ligase connects DNA’s

DNA Helicase

unwinds the double helix of DNA with ATP hydrolysis

SSB proteins

Single Stranded Binding Proteins are added to the unwinded section of DNA to stabilize it

DNA gyrase + example enxyme

Topioisomerase keeps strands from positive supercoiling

DNA Primase and oriC

adds a short sequence of RNA as a primer for the DNA polymerase to start at the origin of replication (oriC)

DNA Polymerase III

catalyzes the addition of deoxynucleotides in the 5’ to 3’ direction

Leading Strand

is catalyzed continuously by the DNA polymerase III, going from the 5’ to 3’ down the DNA strand

Lagging Strand

is catalyzed discontinuously, with Primase adding the primer down the line, and DNA polymerase going “backwards” to add nucleotides in the 5’ to 3’ direction

Created Okazaki fragments

DNA Polymerase I

removed the RNA primer ribonuceleotides, and adds deoxyribonuclotides in its place

DNA Ligase

Joins the deoxynucleotides together after the RNA primer sections are removed

DNA replication is semiconservative

The result of DNA replication is two double helix stands, where 1 strand is “old” and the other strand is “new”.

The parental/old strand is the template strand

Basic Transcription process

RNA polymerase (RNAP) encloses the DNA and opens it up as a transcription bubble

It then catalyzes the addition of ribonucleotides opposite of the template strand, in the 5’ to 3’ direction

As it adds nucleotides, the chain goes out of the RNAP

Parts of Bacteria Transcription

Core Enzyme is the RNA polymerase

Sigma Factors (sigma 70) recognizes the promotor initiation site where transcription begins

Pribnow box

a bacteria promotor sequence that is 10 nucleotides upstream of the start site (-10)

promotor sequence = TTGACA

a bacteria promotor sequence that is 35 nucleotides upstream of the start site (-35)

What are the two Bacterial promotor sequences

Pribnow box = -10 = TATAAT

-35 sequence = TTGACA

Operon

a sequence of genes that encode several enzymes of a particular metabolic pathway

Polycistronic mRNA

FOUND IN Prokaryotes

the mRNA transcribed can be used to translate multiple proteins

Monocistronic mRNA

FOUND IN Prokaryotes and Eukaryotes

the mRNA transcribed can be used to translate a single protein

Bacteria Transcription Termination

Rho-independent vs Rho-dependent

Rho-independent termination

In bacteria

The RNA creates a stem-hoop where the parts touching is a GC-rich inverted repeat, with a Hairpin loop end.

When RNA polymerase reaches the loop, RNA and DNA dissociate

Rho-dependent termination

The Rho protein and RNA polymerase contact each other and cause termination

TATA box

A promoter of 6-8 base pairs, 18-27 nucleotides upstream

Found in archaea and eukarya

TBP

TATA binding protein that recognizes TATA box

BRE

B recognition element is a sequence upstream of the TATA box

Transcription factors that recognize BRE

Archaeal TFB

Eukaryotic TFIIB

Inr

Initiator element at the start that gets recognized by TFB and TFIIB

Basic steps of archeal and eukaryotic transcription

1) TBP binds to the TATA box

2) archeal TRB or eukaryotic TFIIB binds to the BRE

3) RNA polymerase can bind and initiate transcription (same as bacterial)

4) Termination (different between archaeal and eukaryotic)

Eukaryotic transcription termination

5’ gets capped and cut off

Uncapped end gets binded by RNase and degrades it

5’ capping

an methylated guanine’s 5’ end is added to the 5’ end to protect from nucleases (5’ - 5’ linkage)

3’ Polyadenylation

trims the 3’ end, and added the Poly(A) tails, a sequence of 100 Adenines to protect mRNA from nuclease attack

Splicing

the noncoding Introns are removed by spliceosome, and exons are joined together

How are amino acids linked to form polypeptides

By peptide bonds- a carboxyl and the amino nitrogen of two amino acids

Non polar R chains

methyl groups and rings

Polar R chains

OH groups

Acidic R chains

negative Oxygen charge

Basic R chains

Positive nitrogen charge

Primary protein structure

linear sequence of amino acids

Secondary protein structure

hydrogen bonding between O and N of two peptide bonds. form alpha helices and beta sheets

Tertiary protein structure

Disulfide bonds between R groups (cycstine bonds) to form a 3D structure

Quarternary protein structure

many proteins stabilized by disulfide bonds

Structure of tRNA

four leaf clover, single strand, extensive secondary strucure

Function of tRNA

Contains Anticodon sequence to recognize a codon on the mRNA

Linked by aminoacyl-tRNA synthease

Acceptor end of an tRNA

the 3’ end that has an unpaired nucleotide sequence of CCA

Charged tRNA

an amino acid is activated by ATP to form Aminoacyl-AMP

Aminoacyl-AMP is attached to the CCA stem to become “charged”

How many possible codons are there?

64

4 bases, 3 nucleotides (codon)

4³

Open reading frame (ORFs)

a start codon followed by a number of codons and then a stop codon on the same polycistronic mRNA

Degeneracy

a code that lacks one-on-on correspondence between amino acid and codon

Wobble

when some tRNAs can recognize more than one codon, the anticodon forms base pairs with the first two positions, and tolerates irregular pairing on the third position that makes it…

Ribosome subunits in bacteria and archaea

Total is 70 S

30S = 16S rRNA+ 31 proteins

50S

Ribosome subunits in Eukaryotic

total of 80S

40S = 19S rRNA + 33 proteins

60S

what are the ribosomal binding sites

A site, P site, E site

A site

aminoacyl tRNA

where the incoming charged tRNA is attached

P site

peptidyl tRNA

where the growing polypeptide chain is attached to the prior tRNA

E site

exit site

Process of Translation (1-6)

1) tRNA is in the P site to start

2) Incoming tRNA binds to A site

3) Amino acid on the 3’ end of the tRNA in the P site is moved to the amino acid on the A-site (peptide bonds) (elongation factors)

4) tRNA on the P site goes through the E site to exit

5) the tRNA in the A site moves to the P site for the next one

6) Termination occurs when ribosome reaches the stop codon, and Release Factors cleave the polypeptide

Chaperones

catalyze macromolecule folding events to prevent improper folding

uses ATP

Translocases

proteins that transport specific proteins through the membranes

Sec System

transports unfolded proteins into periplasm, OR inserts it into the cytoplasmic membrane

How does the Sec System transport unfolded proteins into the periplasm

Sec A protein binds polypeptides to transport to the channel, where it is translocated by ATP

How does the Sec System insert the translocated protein into membrane?

Uses SRP to bind polypeptide to be inserted using cotranslation

(signal recognition particle)

Tat System

transports folded proteins through cytoplasmic membrane.

the transported proteins contain a short signal sequence that has a pair of arginine residues

How does the Tat system transport?

the TatA protein is the membrane transporter

Binary Fission

rod elongates and then Septum formation splits it in two equal halves

one generation time as occured

Budding Division

the mother cell retains its original identity, and the daughter cell has an unequal cell growth compared to the mother cell

Biofilms

attached polysaccharide matrix containing embedded bacterial cells

Biofilm creation stages

Planktonic (free) cells attach to surface

Sticky matrix is produced

further cell growth and develop leads to biofilm formation

Purpose of Biofilms

to prevent penetration by chemicals, or being washed away.

provides a barrier for cells.

Growth curve stages

Lag, exponential, stationary, decline phases

Lag phase

no growth after inoculation, adjustment period

Exponential phase

cell population doubles at regular intervals

Stationary phase

no net increase or decrease in cell number (some die, some grow)

Decline/ Death Phase

total number of cells decreases due to rapid cell death

Chemostat purpose

is a device for continuous/open system culture

sterile medium is added at the same rate that spent culture medium is removed to keep a steady rate for a specific growth rate

Defined media

know the exact composition of media for specific growth

Complex media

dont know the exact composition for broad growth

Selective medium

inhibits growth of differing mircoorganisms

Differential medium

uses an indicator to differentiate between certain aspects of a mircoorganism family

Total cell counts

uses dried or liquid samples to get cell count

Viable cell counts

uses spread plate or pour-plate method to get alive samples that can grow

Optical density

unit of turbidity at the wavelength specific (OD₅₄₀)

proportional to cell number

Cardinal Temperatures

the minimum, optimum and maximum temperates where growth is either not possible, slow, or the most rapid

Psychophiles

like low temperatures, like 4-15 C

Mesophiles

like midrange temperature (30-45 C)

Thermophiles

like high temperatures (60 C)

Hyperthermophiles

like extremely high temps (106 C)

Molecular adaptions for cold climates (5 of them)

1) enzymes have a-helices = less rigid

2) mole polar bonds

3) more unsaturated fatty acids = less rigid

4) cold shock proteins = chaperones

5) cryoprotectants = antifreeze proteins to prevent ice crystals

Molecular adaptations for hot climates (3 of them)

1) type of amino acid sequence that resists denaturation

2) increases ionic bonds

3) certain phosphates that help stabilize

Neutrophiles

pH of 5.5-7.9

Acidophiles

pH below 5.5

high concentration of protons

Alkaliphiles

pH above 8

bacillus species

water acitivity

a_w

availability of free water

0 is no free water

1 is pure water