Transcript Nucleic acids - Univerzita Karlova v Praze

Nucleic acids
Universal constituents of living matter.
They are concerned with the storage,
transmission and transfer of genetic
information.
Vytášek 2008
Presentation is only for internal purposes of 2nd Medical faculty
Nucleic acids
• polymers in molar mass in the range 20kDa
to 4GDa or more
• two basic types : DNA and RNA
• during last twenty years rapid development
– molecular biology and genetics, genetic
engineering etc.
Components of nucleic acids
• phosphoric acid (H3PO4)
• pentose (5 carbon sugar)
D-ribose
(RNA)
2-deoxy-D-ribose
(DNA)
• heterocyclic bases (nucleobases)
pyrimidine
uracil
(RNA)
thymine
(DNA)
O
NH2
N
N
N
NH
purine
N
N
N
cytosine
NH
adenine
N
HN
H2N
N
NH
guanine
Nucleoside consists of a pentose (ribose or
deoxyribose) linked via a -N-glycosidic bond
to a ring nitrogen of nucleobase.
Adenosine
guanosine
cytidine
Nucleotide
consists of nucleoside which is esterified by phosphoric acid
at either 5´ (usually and exclusively in nucleic acids) or 3´hydroxyl group of pentose moiety. Nucleotide is
monomeric unit of polymeric nucleic acids
(polynucleotides)
• deoxyribonucleotides
– dAMP, dGMP, dTMP
and dCMP (monomers
of DNA)
• ribonucleotides –
AMP, GMP, UMP and
CMP (monomers of
RNA)
AMP
Nucleic acid is polynucleotide chain
which contains mononucleotides linked
by phosphodiester bonds.
• 5´-OH group of nucleotide is joined to 3´OH group of another nucleotide by
phosphodiester bond
• polynucleotide chain contains 5´- and 3´end and by convention nucleic acid
sequence is written from 5´ to 3´ end
RNA
DNA
Most DNAs exist as
double-helix (duplex) structures.
Double-helix (duplex) structure of DNA
• double helix is stabilized by hydrogen
bonds of base pairs (A-T, G-C) but also by
other forces (van der Waals forces, dipoldipol interactions, repulsive forces of
negatively charged phosphoryl groups etc)
• denaturation of DNA involves separation of
complementary strands
• DNA renaturation is the formation of
duplex structure from single strands
The RNA family
Messenger RNA (mRNA)
• transfer of information in a gene (DNA) to
the protein synthetizing machinery
• very heterologous in size and stability, each
molecule is template for specific protein
sequence
• translation of mRNA to protein begins from
5´-terminus
• the most of mRNA molecules contains
polyA tail at the 3´-hydroxyl teminus
(attached 20-250 adenylate residues)
• 5´-end is capped
Transfer RNA (tRNA)
•carries amino acids to the template for protein synthesis
•small molecules (about 75 nucleotides)
•at least 20 species of tRNA in every cell corresponding to each
of 20 amino acids
Ribosomal RNA (rRNA)
• integral component of ribosome (specific
cytoplasmatic structure for proteosynthesis
from mRNA templates)
• the bulk of cellular RNA
• three rRNAs 28S 18S and 5,8S
• catalytic (enzyme like) activity during
creation of peptidyl bond
Small interfering RNAs
(siRNAs)
• around 22 nucleotides in length
• mediate the recently discovered
phenomenon of RNA interference
(RNAi)
• mediate the downregulation of gene
expression - binding to specific mRNAs
labelled them for destruction by
enzymes called endonucleases
MicroRNAs (miRNAs)
• 22–24 nucleotides in length
• downregulate gene expression - binding
to messenger RNAs (mRNAs) causes
preventing mRNAs from being
translated into proteins
Gene silencing in mammals by
microRNAs and small interfering
RNAs
Nature Rev. Genet.
3, 737–747 (2002)
Small nucleolar RNAs
(snoRNAs)
• important for the biosynthesis of rRNAs,
• modify ribosomal RNAs (rRNAs) by
organizing the cleavage of the long prerRNA into its functional subunits (18S,
5.8S and 28S molecules)
Small nuclear RNAs
(snRNAs)
• are constituents of the cellular machinery
(spliceosome) that helps to produce
mRNA
• removing the non-coding regions (introns)
of genes and piecing together the coding
regions (exons) to be translated into
proteins
• some of these snRNAs have been shown
to be the functional enzymes in the
splicing reaction
Ribozyme
• an RNA molecule that catalyzes a chemical
reaction
• from ribonucleic acid enzyme (called also
RNA enzyme)
• many natural ribozymes catalyze either the
hydrolysis of one of their own phosphodiester
bonds or the hydrolysis of bonds in other
RNA
Chemical differences between
DNA and RNA
carbohydrate
pyrimidine base
structure
stability at high pH
DNA
deoxyribose
thymin
double-helix
resistent
RNA
ribose
uracil
single helix
cleaved
Estimation of nucleic acids
• content of phosphorus (bulk of pure nucleic
acids)
• content of bases (absorption of UV light)
• content of pentose
• estimation of native chains by intercalating
stains
Ethidium bromide intercalation
Intercalation induces structural
distortions