讲义生化课件f2nucleic acids structure

1、1 Nucleotides and nucleic acids 2 Nucleic acids structure 3 DNA organization in chromosomes 4 DNA replication 1 The primary structure of nucleic acids 2 The ABZs of DNA Secondary Structure 3 Secondary and Tertiary Structure of RNA 4 Denaturation and Renaturation of DNA 5 Nucleic Acid Hybridization T

2、he sequential order of nucleotides in a polynucleotide, is the so-called primary structure of nucleic acids. Two basic protocols for nucleic acid sequencing are in widespread use: the chain termination or dideoxy method of F. Sanger and the base-specific chemical cleavage method developed by A. M. M

3、axam and W. Gilbert. Double-stranded DNA molecules assume one of three secondary structures, termed A, B, and Z. Fundamentally, double-stranded DNA is a regular two chain structure with hydrogen bonds formed between opposing bases on the two chains. the four bases commonly found in DNA (A, C, G, and

4、 T) do not occur in equimolar amounts and that the relative amounts of each vary from species to species. The ratios of adenine to thymine and of guanine to cytosine were found to be nearly 1.0 in DNA samples from all species studied. The number of pyrimidine residues always equals the number of pur

5、ine residues. These findings are known as Chargaffs rules: A = T; C = G; pyrimidines = purines. James Watson Francis Crick James Watson and Francis Crick, working in the Cavendish Laboratory at Cambridge University in 1953, took advantage of Chargaffs results and the data obtained by Rosalind Frankl

6、in and Maurice Wilkins in X-ray diffraction studies on the structure of DNA to conclude that DNA was a complementary double helix. The fact that the DNA molecule is composed of bases, deoxyriboses, and phosphate groups linked together as a polydeoxyribonucleotide. X-ray diffraction pattern of DNA fi

7、bers, suggesting a helical structure with two distinctive regularities of 3.4 and 34 Angstroms along the axis of the molecule. Chargaffs discovery on the quantitative relationships between the bases (A=T, G=C). The DNA molecule is a right-handed double helix containing two antiparallel strands. The

8、phosphate-deoxyribose backbones are on the outside of the helix (forming a “hydrophilic surface”), whereas the purine and pyrimidine bases are stacked inside (the base-stacking interactions make a major nonspecific contribution to the stability of the duplex). 34 The planes of the bases are perpendi

9、cular to the helix and the planes of the deoxyribose rings are nearly at right angles to those of the bases. The two antiparallel chains are complementary to each other through hydrogen bonds between pairs of bases. Adenine is always paired with thymine (with two H-bonds), guanine with cytosine (wit

10、h three H-bonds). The diameter of the proposed helix is about 20 , adjacent bases are separated by 3.4 and related by a rotation of about 36 with the helical structure repeats about every 10 residues on each chain at intervals of about 34 . The DNA molecule contains two kinds of grooves, a major gro

11、ove and a minor groove. The major groove display more distinctive potential H-bonding features than the minor groove. The double-helical model of DNA immediately suggested a mechanism for the replication of DNA. Genetic information has to be replicated (duplicated). The double helix model for DNA is

12、, in effect, a pair of templates, each of which is complementary to the other. It was proposed that at replication, the parent strands become separated (H-bonds are broken), and each forms the template for biosynthesis of a complementary daughter strand. Watson, Crick, and Wilkins shared the Nobel P

13、rize in medicine or physiology in 1962 for this brilliant accomplishment. The discovery of the DNA double helix revolutionized biology: it led the way to an understanding of gene function in molecular terms (their work is recognized to mark the beginning of molecular biology). DNA is remarkably flex

14、ible molecule with many rotatable bonds. The duplex structure proposed by Watson and Crick is referred as the B-form DNA, and is found to be the most stable structure for a random nucleotide sequence under physiological conditions. Thus it is the standard structure for DNA molecules. At reduced humi

15、dity the DNA molecule will take the A-form: it is still a right-handed duplex made up of antiparallel strands held together by Watson-Crick base pairing. The A-form helix is wider and shorter than the B- form helix. The plane of the base pairs in A-DNA is tilted about 20 with respect to the helix ax

16、is. The Z-form DNA is a left-handed double helix in which backbone phosphates zigzag. The Z-form DNA is adopted by short oligonucleotides that have sequences of alternating pyrimidines and purines (e.g., CGCGCG). The biological roles of Z-DNA is uncertain (may play roles in gene expression and genet

17、ic recombination). Intact DNA molecules from bacteria, some viruses, mitochondria, and chloroplasts are circular and supercoiled. The axis of the double helix can be twisted to form a superhelix (the circular DNA without any superhelical turns is known as a relaxed molecule). Supercoiling makes the

18、DNA molecule more compact thus important for its packaging in cells. RNA molecules do not form simple, regular secondary structure but many form complex and unique three dimensional structures. (G1). Single strand RNA tends to take right-handed helical conformation. Self-complementary sequences on a

19、 RNA molecule lead to specific structures. The standard base-pairing rules are followed (i.e., A with U, G with C). Intrastrand base pairing makes structures including bulges, internal loops, and hairpins (helical). Messenger RNA (mRNA) serves to carry the information or “message” that is encoded in

20、 genes to the sites of protein synthesis in the cell, where this information is translated into a polypeptide sequence. Messenger RNA is synthesized during transcription, an enzymatic process in which an RNA copy is made of the sequence of bases along one strand of DNA. In prokaryotes, a single mRNA

21、 may contain the information for the synthesis of several polypeptide chains within its nucleotide sequence.- Polycistronic In contrast, eukaryotic mRNAs encode only one polypeptide. -Monocistronic, eukaryotic mRNAs are synthesized in the nucleus in the form of much larger precursor molecules called

22、 heterogeneous nuclear RNA, or hnRNA. hnRNA molecules contain stretches of nucleotide sequence that have no protein-coding capacity. These noncoding regions are called intervening sequences or introns because they intervene between coding regions, which are called exons. Introns must be spliced out

23、before the message can be translated. Eukaryotic hnRNA and mRNA molecules have a run of 100 to 200 adenylic acid residues attached at their 3-ends, so-called poly(A) tails. This polyadenylylation occurs after transcription has been completed and is believed to contribute to mRNA stability. Transfer

24、RNA (tRNA) serves as a carrier of amino acid residues for protein synthesis. Transfer RNA molecules also fold into a characteristic secondary cloverleaf. Each cloverleaf consists of four H-bonded segmentsthree loops and the stem where the 3- and 5-ends of the molecule meet. These four segments are d

25、esignated the acceptor stem, the D loop, the anticodon loop, and the TC loop. Extra or Variable loop Transfer RNA All tRNA molecules possess a 3-terminal nucleotide sequence that reads CCA. The amino acid is attached as an aminoacyl ester to the free 3-OH of the terminal A residue. Amino acid accept

26、or stem D loop T loop Anticodon loop Variable arm Cloverleaf secondary structure of tRNA The amino acid acceptor stem is at one end of the L, the anticodon at the opposite end of the L. The D and T C loops form the corner of the L. Primary tRNA transcripts undergo a series of posttranscriptional pro

27、cessing The extra sequences at the 5 and 3 ends are removed by RNase P and RNase D respectively. The CCA sequence is generated at the 3 end. Some of the bases in tRNA molecules are modified by methylation, deamination, reduction and others. Ribosomes, the protein-synthesizing machinery of cells, are

28、 composed of two subunits, called small and large, and ribosomal RNAs are integral components of these subunits. Ribosomes The proposed secondary structure for E. coli 16S rRNA The 16S, 23S and 5S rRNAs in bacteria are all generated from a single 30S pre-rRNA (about 6.5 kb, transcribed by RNA polyme

29、rase). The 18S, 28S and 5.8S rRNAs in eukaryotes are generated from a single 40S pre-rRNA (14 kb). The 5S rRNA in eukaryotic cells is generated separately (transcribed by RNA polymerase III). The 18S, 5.8S, and 28S rRNAs in eukaryotic cells are derived from one pre-rRNA molecule (the processing need

30、s small nucleolar RNA-containing proteins). The M1 RNA in ribonuclease P is catalytic The intron in the pre-rRNA of Tetrahemena is self-spliced Duplex nucleic acids unwind to form two single strands at extreme pH and high temperature with changed physical properties. Viscosity decreases sharply. UV

31、absorption at 260 nm increases significantly, an effect called hyperchromism or hyperchromic effect. (base stacking decreases absorption). hypochromic effect The rise in absorbance coincides with strand separation, and the midpoint of the absorbance increase is termed the melting temperature, Tm . (

32、half of the duplex chain is separated) The unwinding (i.e., denaturation) of the double helix is called melting because it occurs abruptly at a certain temperature. Each species of DNA has a characteristic melting temperature (Tm or tm) . Tm of a DNA molecule depends markedly on its base composition: DNA with higher content of GC base pairs has higher Tm because there are three H-bonds between each GC base pair, but only two H-bonds between the AT pair (Tm has an approximate linear relationship with (G+C)%). DNA segments rich in AT base pairs are melted first (at low