Genes are composed of nucleic acids (usually DNA)

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Genes are composed of nucleic acids (usually DNA) • Pneumococcus can be transformed from an avirulent to a virulent strain • DNA is the transforming principle • DNA in bacteriophage particles appears in the progeny, but very little protein does.

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Genes are composed of nucleic acids (usually DNA). Pneumococcus can be transformed from an avirulent to a virulent strain DNA is the transforming principle DNA in bacteriophage particles appears in the progeny, but very little protein does. Structures of nucleic acids. Nucleotides - PowerPoint PPT Presentation

Transcript of Genes are composed of nucleic acids (usually DNA)

Page 1: Genes are composed of nucleic acids (usually DNA)

Genes are composed of nucleic acids (usually DNA)

• Pneumococcus can be transformed from an avirulent to a virulent strain

• DNA is the transforming principle

• DNA in bacteriophage particles appears in the progeny, but very little protein does.

Page 2: Genes are composed of nucleic acids (usually DNA)

Structures of nucleic acids

Nucleotides

DNA structures

Sedimentation and Electrophoresis

Page 3: Genes are composed of nucleic acids (usually DNA)

A simple view of DNA

AGCCTCGCATTCGGAGCGTA

Page 4: Genes are composed of nucleic acids (usually DNA)

Nucleotides

• 3 components to nucleotides:

– Purine or pyrimidine base

– Ribose (RNA) or 2-deoxyribose (DNA) sugar

– Phosphate

• Base + sugar = Nucleoside

• Base + sugar + phosphate = Nucleotide

Page 5: Genes are composed of nucleic acids (usually DNA)

Types of bases in nucleotides

Pyrimidine

N

N

H

O

NH2

Cytosine

N

HN

H

O

O

Uracil

N

HN

H

O

O

CH3

Thymine

Amino- Keto-

Page 6: Genes are composed of nucleic acids (usually DNA)

Nucleotides: purine bases

N

N N

N

H

HN

N N

N

H

NH2

H2N

O

Adenine Guanine

6-aminopurine A keto-purine

Page 7: Genes are composed of nucleic acids (usually DNA)

Bases are attached to C1’ of the sugar via an N-glycosidic bond

N

N N

N

NH2

O

OH

CH2HO

Adenosine

3'

5'1'

2’-deoxy- , a nucleoside

Page 8: Genes are composed of nucleic acids (usually DNA)

Phosphate is attached to C5’ of the sugar

1st phosphate is a phosphoester, others are attached as phosphoanhydrides.

-O-P-

O=

O

NTP is

-O-P-

O=

O -O-P-O

O=

O

Obase

OH OHphosphoanhydride

phosphoester

-

Page 9: Genes are composed of nucleic acids (usually DNA)

Structure of a dinucleotide

The 3’ C of one nucleotide is linked to the 5’ C of the next nucleotide in a phosphodiester linkage.

O

O

CH2O

3'

POO

O

HN

NN

N

NH2

O

OH

CH2OPO

O

H

N

N

O

NH2

5' cytidylyladenylateor5'pCpA

5'

3'

5'

Page 10: Genes are composed of nucleic acids (usually DNA)

Nucleic acids are linear chains of nucleotides

• The 3’ C of one nucleotide is linked to the 5’ C of the next nucleotide.

• The linkage is by a phosphoester.

• The chain has an orientation defined by the sugar-phosphage backbone.

• One terminal nucleotide has a “free” 5’ end, and the other has a “free” 3’ end.

• Thus we designate orientation by 5’ to 3’.

Page 11: Genes are composed of nucleic acids (usually DNA)

More on orientation of chains of nucleic acids

• 5’ ACTG 3’ is different from 3’ ACTG 5’

• Unless specified otherwise, a chain is written with the 5’ end on the left and the 3’ end on the right.

• When complementary strands in DNA are written, usually the top strand is written 5’ to 3’, left to right, and the bottom strand is written 3’ to 5’, left to right.

5’ GATTCGTACCG

3’ CTAAGCATGGC

Page 12: Genes are composed of nucleic acids (usually DNA)

Basics of DNA structure

• 2 complementary strands of nucleic acids

• Complementarity is based on H-bonding between– Keto bases with amino bases– Pyrimidines with purines

• A pairs with T (or U)

• G pairs with C

• The complementary strands are antiparallel.

• The complementary strands are coiled around each other.

Page 13: Genes are composed of nucleic acids (usually DNA)

Duplex DNA• Two strands coil around each other.• Right-handed coils (B form and A.• Coils form major and minor grooves.• Strands have opposite polarity (antiparallel).• Opposing bases in strands are complementary.• Different edges of paired bases are exposed in major and

minor grooves.• Sugar-phosphate backbone is on the outside, bases on the

inside– B-form DNA: base pairs are close to center of long axis of the duplex.– A-form nucleic acids: base pairs stack away from long axis.

Page 14: Genes are composed of nucleic acids (usually DNA)

Implications of complementarity

• One chain (strand) of DNA can serve as the template for synthesis of the complementary chain.

• DNA replication: sequence of nucleotides in one chain of the duplex determines the sequence of nucleotides in the other chain.

• Transcription: sequence of nucleotides in one chain of the duplex determines the sequence of nucleotides in mRNA or its precursor.

Page 15: Genes are composed of nucleic acids (usually DNA)

Base pairs in DNA

NN N

N

O

O

N H

H

N

HH

H

deoxy-ribose deoxy-

ribose

N

N N

N

NN N

N

O

NH

H

H

O CH3

deoxy-ribose deoxy-

ribose

Guanine : Cytosine Adenine : Thymine

Major groove

Minor groove

Major groove

Minor groove

Page 16: Genes are composed of nucleic acids (usually DNA)

Major types of duplex nucleic acid structures

• B form– Most common form of DNA– Right handed DNA-DNA helix– Base pairs stack close to DNA central axis

• A form– right handed RNA-DNA and RNA-RNA helix– Base pairs stack away from the central axis

• Z form DNA– Repeating purines and pyrimidines– Left-handed helix–May serve as some regulatory signal in cells

Page 17: Genes are composed of nucleic acids (usually DNA)

Forms of nucleic acid duplexes

B-form DNA A-form (e.g. duplex RNA)

Z DNA

Page 18: Genes are composed of nucleic acids (usually DNA)

Helical parameters for B, A and Z nucleic acids

B A Zhelix sense RH RH LHbp per turn 10 11 12vertical rise per bp 3.4 2.56 3.7 Angstromsrotation per bp +36 +33 -30 degreeshelical diameter 19 23 18 Angstroms

Page 19: Genes are composed of nucleic acids (usually DNA)

Hyperchromic shift when DNA is denatured

Native duplex DNA Denatured, strands are separate

+

denaturation by heat orincreasing pH

renaturation by cooling or lowering pH

lower A260

higher A260

hyperchromic

hypochromic

1.0

1.2

1.4

Tm= melting temperature, midpoint of the transition

Temperature

A260

Page 20: Genes are composed of nucleic acids (usually DNA)

Factors that affect melting temperature, p. 85

• The melting tempera-ture (Tm) increases as– Increase G+C– Increase ionic strength

(or )

• Tm decreases as– Increase denaturants– Increase number of

mismatches

Tm = 0.41 (% G+C) + 16.6 log M + 81.5 -0.7 (% formamide) -1o (% mismatch)

% G+C

Tm

= 0.06 = 0.21

Page 21: Genes are composed of nucleic acids (usually DNA)

Electrophoresis to measure SIZE

For molecules of the same shape, logM is inversely proportional to d.

For molecules of the same size, more compact forms, such as supercoiled DNA, moves faster than more extended forms, such as linear DNA.

+

-

Size markersDNA samples

Page 22: Genes are composed of nucleic acids (usually DNA)

Example of gel electrophoresis

Alpha-globingenePCR product217 bp

Markers

100

200300400 base pairs