The Basics: A general review of molecular biology: DNA Transcription RNA Translation Proteins

DNA (deoxy-ribonucleic acid) is the genetic material -think of it as the blueprint

DNA structure-- a polymer of nucleotides

Nucleotides have :

2) a ring-shaped nitrogen base

1) a 5 carbon sugar 3) a phosphate group Fig 0-3

Things to notice about the sugar: --sugars can circularize by eliminating and H20 molecule and forming a bond between hydroxyl groups --the carbons in the sugar are given numbers in standard Nomenclature, designated as prime to distinguish from carbons on the nitrogen base

these numbers are used to distinguish critical sites in The nucleotide and in the DNA strand.
ribose is a 5-carbon sugar Fig 0-1

2-deoxy-ribose is different from ribose in that it lacks a hydroxyl group (-OH) on the 2 carbon

DNA-- 2-deoxy-ribose RNA--ribose

The chemical difference associated contributes signicantly to the differences between DNA and RNA biochemistry

Fig 0-1

The OH groups on the 5 and 3 carbons are the reactive groups through which nucleotides become joined

Fig 0-1

The OH groups on the 5 and 3 carbons are the reactive groups through which nucleotides become joined a nucleotide includes one phosphate group joined at the 5 position.. PO4

There are 4 nitrogen bases in DNA:

NH 2 C N1 C H
2 6 5 4

O C C N
7 8

H C C N C N H H H H C4 N3 C
5 6

C H3 H C N H NH 2 C N C O Th y m i n e
1

H C N H H2N C

C C N

N H

C O C y t os i n e

Ad e n i n e

Gu a n in e

Adenine and Guanine have 2 rings--purines Cytosine and Thymine have 1 ring--pyrimidines Fig 0-2

PO4 What to know about the phosphate: 1) linked at 5 carbon 2) can have 1, 2, or 3 phosphate residues (nucleotide mono-phosphate, nucleotide di-phosphate , nucleotide tri-phosphate) 3) ***the oxygens of the phosphate group are negatively charged at physiological pH. Therefore DNA carries a large net negative charge!

Fig 0-3

The polynucleotide chain

To form the polynucleotide chain, the oxygen of the 3' hydroxyl group on the chain attacks the phosphate of a nucleotide triphosphate eliminating H2O and releasing the two outermost phosphate residues.
Bond formation:
base base
OO P O O OP O O OP O
3' 5'

OO P O O

OP O O

O5'

CH2

P O

CH2

sugar
X

sugar
X

..
O-

OH

O 3'

base
OO P O O OP O O
5'

P O

base
5'

The phosphodiester bond

P O

CH2

sugar
X

CH2

sugar
OH X

3'

OH

3'

*******Notice that the DNA chain is synthesized in a 5 to 3 direction. Fig 0-4

5 end ***There is an asymmetry to the DNA chain!

5 end--phosphate group 3 end--free hydroxyl group

Read as:
3 end

5ATGC 3

Fig 0-5

DNA is double-stranded--two polynucleotide chains Hydrogen bonds between bases hold these together
0 5

Guanine

Cytosine

Adenine

Thymine

G and C have 3 H-bonds

A and T make 2 H-bonds Fig 0-6

DNA strands are arranged in an anti-parallel manner 5 3

Complementary
(Not complimentary)

Fig 0-7

Critical Properties of DNA 1) Negative charge (will move toward a + electrode!) 2) DNA can be denatured and renatured (nucleic acid Hybridization). 3) DNA is soluble in water. 4) DNA is insoluble in ethanol. 5) DNA absorbs UV light. 6) DNA can be stained and amounts of DNA can be Measured using ethidium bromide. Ch. 0-5

UV absorption properties of DNA Bases absorb UV light with a max absorbance at 260 nm The amount of light absorbed is proportional to the amount of DNA in solution

1.0 O.D.

50g/l DNA concentration

100g/l

An A260 reading of 1.0 corresponds to a 0.05 mg/ml concentration of DNA (by denition) Ch. 0-5

Ethidium bromide intercalates into the DNA double helix EthBR uoresces under UV light, enabling us to see DNA

no uorescent color

uorescent
Fig 0-8

Ethidium bromide intercalates into the DNA double helix EthBR uoresces under UV light, enabling us to see DNA
!#*&^#%**#@! degenerins

des(bz29)

EthBR can intercalate degenerative death DNA! Be very into your pathway??? cautious in lab use!

Genes, chromosomes and genomes

Gene: DNA devoted to making one specic polypeptide Genes are housed on chromosomes All DNA of an organism makes up its genome Ch. 0-6

Central dogma

DNA
transcription

RNA
translation

Protein

Ch. 0-6

Transcription
--DNA is rst transcribed into mRNA before protein is made --Product is messenger RNA or transcript --Process is transcription

Transcripts are made of single-stranded RNA

Remember, RNA is fundamentally different from DNA: Uracil rather than thymine is used (can pair with A) Ribose is the sugar mRNA is predominantly single-stranded

Fig 0-9

RNA polymerase is the enzyme that catalyses mRNA synthesis The chain is extended from the 3 end The chain grows in the 5 to 3 direction

Fig 0-10

The DNA strand copied is called the coding strand

Fig 0-11

The promoter is the site where RNA polymerase binds to initiate transcription

Promoters are part of the DNA of the gene (called the 5 non-coding region) but are not included in the transcript itself

Fig 0-12

Eukaryotic messages are processed

1) 5 cap 2) 3 poly (A) tail 3) splicing-exons stay in the message;introns are clipped out

Fig 0-12

Protein synthesis: translation (mRNA to protein)

Proteins have catalytic and structural functions Proteins with catalytic functions are enzymes

Amino acids are the building blocks of proteins: Common to all: Hydrogen atom

Amino group

Carboxyl group

Distinguishing feature: R group; side chain conveys specic chemical properties

Different amino acid chains have different chemical properties

Fig 0-13

Protein synthesis--chain elongation mechanism

Fig 0-14

Proteins also have a polarity to them-distinct beginning and end

Free amino group At the rst residue Free carboxy group at the last residue

OH

Fig 0-15

The Genetic Code

Codon--mRNA triplet that signals single AA addition

1) triplet--3 bases in a row code specic amino acids 2) code is degenerate 3) initiation codons start (AUG = Met) 4) stop codons terminate (UAA, UAG, UGA)

Fig 0-16

5) frame is critical AAUUCGAGUUUGUG ASN-TRP-SER-LEU ILE-ALA-VAL-CYS PHE-GLU-PHE-TRP

1 2 3

Reading frame is set by the initiation of translation

Fig 0-17

Translation: mRNA to protein

Transfer RNA (tRNA) are the adaptors

complementary to codon

Translation on the ribosome

Fig 0-18

Regulation of gene expression and protein activity

Fig 0-19