Molecular Bioscience Exam 1 (2)

Replication Directionality

Bidirectional having two replication forks

Stop Codon

Termination of protein synthesis coded in mRNA

ORC

Binds DNA at origin replication

Helicase

Unwinds DNA at the replication fork by breaking H bonds

Topoisomerase

Cut DNA backbones to remove knots and other entanglements

Single-Stranded Binding Proteins

Stabilize single stranded DNA, preventing reformation of double stranded DNA

DNA pol III

Synthesize new DNA in leading and lagging strand

RNase H

Removes RNA primer

RNA Primer

Acts as start point for DNA replication

DNA Primase

Creates Primer

Telomerase RiboProtein Complex

Resolves the end replication problem by extending telomeres in the 3’ end acting as a template

Telomerase

Reverse transcriptase is able to create DNA from RNA template

Telomerase RNA Component (TERC)

Provides RNA template for adding repeats at the 3’ end of chromosomes

DNA Damage

Changes in DNA structure (abnormality), mismatched bases, damaged bases, changes in bonds.

Nucleotide Excision Repair

Repairs UV-light chemical damage

Base Excision Repair (BER)

Repairs oxygen radicals, hydrolysis, alkylating agents

Mismatch Repair (MMR)

Repairs replication errors (mismatch, insertion, deletion).

DNA Mutations

Change in DNA sequence

increased DNA damage

Increased DNA mutation

Gene

Basic unit of heredity, a linear sequence of nucleotides along a segment of DNA, providing instructions for the synthesis of RNA (protein coding)

Transcription Bubble

The region of DNA where the double helix unwinds, allowing the enzyme RNA polymerase to access the DNA bases and synthesize a complementary RNA strand during transcription

Transcription

Only uses one strand of DNA and produces RNA, in one direction for the specific gene.

TF Binding Site

Area where transcription factors bind to regulate gene expression

Promoter Region

Upstream to gene, binding site for RNA pol and TF, allowing transcription

Terminator Region

End of transcription of gene

mRNA

Code for proteins

rRNA

Ribosomal, form the basic structure of the ribosome and catalyze protein synthesis

tRNA

Central to protein synthesis as adaptors between mRNA and amino acids

siRNA

Turn off gene expression by directional degradation of selective mRNAs and the establishment of compact chromatin structures.

miRNA

Regulate gene expression typically by blocking translation of selective mRNAs

Pre-mRNA Processing

Capping, Splicing, and Poly-A Tail in the nucleus

Splicing

Introns removed

Capping

5’-methyl G Cap protecting mRNA from degradation

Tail

3’ Poly A tail, protects from degradation and allows for movement outside of nucleus

mRNA Splicing

Removing introns and ligating exons using the spliceosome with conserved sequences at 5’ and 3’ splice sites as well as branch site (YURAC)

Function fo Smaller RNA in Spliceosome

Recognize conserved sequence in introns by base pairing

Transcription Factors

Regulate the rate of transcription by binding the regulatory DNA sequence (TF binding site). Modular protein that has DNA binding domain, and one or two domains for activation or repression

Transcription Activator

Transcription factor that promotes gene transcription

Transcription Repressor

Transcription factor that inhibits gene transcription

TF Activators

Open Chromatin Structure

TF Repressors

Highly compact chromatin structure

Peptidoglycan

Bacterial wall

Penicillin

Binds and inactivates enzymes that crosslink peptidoglycan of bacterial cell wall

Ribosome Scan Model

Ribosome binds to 5’ cap and scan the first AUG, if its good it will start protein synthesis, if bad it will not.

Energy Production from FA

Beta Oxidation + TCA Cycle

Net Yield of FA Energy

107 ATP per 16C FA

RT-PCR

Use reverse transcriptase (RT) to turn RNA into single-stranded DNA, then regular PCR.

Real-Time PCR (qPCR)

PCR combined with dsDNA binding dye and a camera. The camera determines the amount of PCR product after each PCR cycle. The machine tells you how many cycles of PCR are needed for the signal to reach a specific level (threshold). Lower PCR cycle means more DNA in the sample.

PCR (Polymerase Chain Rxn)

Uses a DNA primer pair and a DNA-dependent pol to amplify a specific DNA sequence in test tube (DNA replication)

Western Blot

Protein samples are treated with SDS and heated so all proteins are (-) and denatured. Run proteins on a gel to separate them based on size, and uses a specific antibody to recognize the protein of interest. Allows for a specific protein to be visualized and analyzed.

Northern Blots

Detect RNA based on hybridization. RNA is heated/denatured before running on a gel to destroy secondary and tertiary structure. RNA separated based on size, then transferred to (+) charged membrane for hybridization.

Southern Blots

Detects specific DNA based on specific base pairing between a DNA or RNA probe and a complementary unlabeled DNA.

Hybridization

Radio-labeled denatured DNA or RNA probe to mesh with denatured DNA on the membrane, based on base pairing.

Electrophoresis

Separation of charged molecules in an electric field based on size (not charge). DNA or RNA loaded in (-) side and smaller molecules move towards (+) faster.

Accurate Translation of mRNA

Requires aminoacyl tRNA-synthetase and base pairing of mRNA codon/tRNA anticodon.

Aminoacyl tRNA-synthetase

enzymes that ensure accurate protein synthesis by catalyzing the attachment of the correct amino acid to its corresponding transfer RNA

tRNA

Brings activated AA to ribosome based on base-pairing of mRNA codon with anticodon

Flouride

Inhibits enolase, inhibiting glycolysis, increasing growth of cariogenic bacteria, and decreasing demineralization of enamel.

Rifampicin

Inhibits initiation of transcription in bacteria by inactivating RNA pol. Remains bound to promoter, blocking initiation

Actinomycin D

Binds tightly and specifically to double-helical DNA and inhibits both transcription and DNA replication (interferes with RNA and DNA pol). Inhibit both bacteria and eukaryotes

alpha-Amantinin

Inhibits mRNA synthesis of eukaryotes but not bacteria by binding RNA pol II, found in poisonous mushrooms.

Fat-Soluble Vitamins

A, D, E, and K (storage)

H2O Soluble Vitamins

B-Complex and C (no storage)

Regulation of Gene-Specific TF Activity

Specific cell types express a specific set of TFs. Even when expressed, TFs are inactive, requiring extracellular signals to regulate the activity of transcription factors. Function of TF can be lost

AD

Protein domain in TF interacting with other proteins or protein complexes in transcription machinery (RNA pol) enhancing recruitment of RNA pol II, or changing chromatin structure (euk)

ETC Inhibitors

1. Decrease O2 consumption

2. Increasing NADH/NAD+ and FADH2/FAD ratio

3. Decreasing ATP production

PTH in Kidney

Stimulates renal tubular Ca2+ reabsorption, stimultes synthesis of 1,25(OH)2D3, increases Ca2+ absorption in small intestine

Calcitonin in Kidney

Inhibits renal tubular Ca2+ reabsorption

PTH in Bone

Function together to mobilize Ca2+ from previously formed bone by stimulating bone resorption by osteoclasts, making it available extracellularly

Calcitonin in Bone

Directly inhibits osteoclasts, enhances osteoblasts

Calcitonin

Increase Ca2+ deposition in bones, decrease Ca2+ uptake in intestines, decrease Ca2+ reabsorption from urine.

Parathyroid Hormone

Increases Ca2+ releases, increase ca2+ uptake in intestines, and increase ca2+ reabsorption from urine

Calcium-Phosphate Metabolism

Parathyroid Hormone, Vitamin D, Calcitonin

Intestine, Bone, Kidney

Vitamins

Fat vs water soluble

Nutrient Classes

Water, protein, carbs, fats, minerals, vitamins (water most important)

GLUT-2 Processes

Liver up glucose to make glycogen and FA, pancreatic beta cells take up glucose to activate insulin release/production

Glucagon

Low blood glucose increases the release of this hormone from alpha cells of the pancreas (increases blood glucose levels and increasing energy supply)

Blood Glucose Regulation

Gluconeogenesis, glycogen synthesis, glycogen breakdown as well as FA synthesis

GLUT2

Functions at high glucose levels (high blood glucose sensor)

GLUT4

Insulin-stimulated glucose uptake; stimulated by exercise in skeletal muscle

GLUT1/GLUT3

Responsible for tissues to bring in glucose from blood to use for energy.

Erythrocytes (RBC)

Only use glucose (glycolysis) as source of energy

Carbons from FA

It can only make acetyl-CoA (succinyl-CoA product of degradation of odd-chain fatty acids)

Carbons from AA

Make glucose, FA , and base, and TCA intermediates, but mainly used for protein synthesis (except leucine and lysine)

Carbons from Glucose

Able to make FA, AAs, and bases, as well as intermediates in TCA (increasing efficiency)

AA in ATP Generation

Primarily used for protein synthesis and bases, not for energy production (but still possible)

Sugar ATP Generation

All cells use this for ATP, sole/main ATP source for RBC and neurons (w/o oxygen)

Fatty Acid ATP Generation

Main energy source for most cell types, produce ketones under starving

ATP Generation

From FA, glucose, AA, NOT nucleic acids