DNA dan RNA UNIT STRUKTURAL DALAM SEL Agustina Setiawati, M.Sc., Apt

Albert, 1994, Molecular Biology of the Cell, Mc Graw Hill REFERENSI

Nucleus Chromosome Kromosom dalam sel Each cell contains DNA. DNA is packaged into compact units called chromosomes. Basically a chromosome is one long chain of genetic material. It is the microscopic, rod-shaped, threadlike part of the cell that carries hereditary information in the form of genes.  Every cell has chromosomes and all individuals in the same species have the same number of chromosomes.  Within an individual, every cell has the same number of chromosomes and generally come in pairs (except in sex cells). People with Down Syndrome have 3 copies of chromosome 21. telomeres, these molecular chains have often been compared to the blank leaders on film and recording tape. Indeed, telomeres seem to perform a similar function in aligning the DNA molecule during the replication process. Protecting the vital DNA molecule from being copied out of synch, these telomeres provide a kind of buffer zone where asynchronous replication errors (that are inevitable) will not result in any of the vital DNA sequences being lost. As a cell gets older, it is under attack by oxides and other so-called free-radical chemicals in the body and environment. We survive as living beings because our cells have the ability to duplicate themselves before being killed by these natural causes. Each time our cells duplicate themselves, a small portion of the DNA molecule is lost and not copied. The loss is usually to the telomere and so the effect is usually insignificant. Scientists recently noted that the length of these telomere chains were shorter as we grew older. Eventually, the telomeres become so shortened that the losses in replication begin to effect the vital DNA molecule sequence and prevent the cell from being able to duplicate itself. This is why we age. Chromosome

Kromosom tanpa histon

Each cell has 46 chromosomes (in 23 pairs) ~30,000 genes are arranged along the 23 types of chromosomes

Genes come in homologous pairs The location of a gene on a chromosome = locus. Alternative forms of genes that influence a characteristic = alleles.

A gene pair of two same alleles (i.e., AA) = homozygous A gene pair of two different alleles (i.e., AO) = heterozygous. DNA Introduction, cont.

Genotype: The particular combination of genes present in the cells of an individual (AA or AO) Phenotype: The physical trait such as, shape, color, blood type, etc. DNA Introduction, cont.

Masing2 single linier DNA Human 4 x 109  22 autosom yang berbeda 2 sex kromosom Masing2 single linier DNA E. Coli 4.7 x 106  1 Chromosom 40 – 50 loop superkoil

Kromatin Merupakan material inti yg mengandung kode genetik Kode genetik tsb tersimpan dlm kromosom Bdsk daya serapnya thdp lar pewarna dibedakan mjd 2 : heterokromatin (menyerap wrn dg kuat) & eukromatin (kurang kuat menyerap wrn) Heterokromatin merup btk rapat (condensed) dr kromatin, shg terlihat spt noda yg rapat. Bnyk terlihat saat sel istirahat, spt limfosit atau sel memori yg menanti antigen asing. Heterokromatin inaktif dlm transkripsi Eukromatin bbtk halus, bnyk tdpt dlm keadaan aktif (sel aktif melakukan transkripsi).

Kromatin Berdasarkan lokasinya kromatin dibedakan mjd 3 daerah : Kromatin perinukleolar, berada di sekeliling nukleolus Kromatin internukleolar, berada dlm nukleolus Kromatin periferal, berikatan dg selaput sel. Kromatin nukleolar & periferal merup heterokromatin

Kromatin Ditinjau dr peranannya sbg materi genetik, heterokromatin dibagi mjd 2 : heterokromatin fakultatif & heterokromatin konstitutif. Heterokromatin konstitutif selamanya tdk aktif & tetap dlm keadaan mampat selama daur hdp sel Heterokromatin fakultatif tdk selamanya berada dlm keadaan mampat. Pd saat ttt secara ajeg kromatin ini terurai & saat terurai mrk dpt disalin Dr analisis kimia ternyata kromatin tdr dr DNA, RNA & protein. Protein yg tdpt di kromatin tdr dr 2 jenis : histon & non-histon

PROTEIN HISTON Histon merupakan protein bersifat sangat basa, yg disebabkan oleh adanya asam amino lisin & arginin dlm jml cukup bnyk. Protein non-histon tdpt lbh dr bbrts jns, bervariasi sesuai dg jenis selnya, mis. Aktin, tubulin, RNA polimerase, asetil transferase dll 146 pasang basa DNA, 1 oktamer histon. 8 buah histon mbtk oktamer yg tdr dr 4 psg masing2 : H2A, H2B, H3 & H4 Set 8 histon tsb dsbt nuklosome, 10 nm fibril nukleoprotein. Nukleosome dipisahkan oleh daerah antara 4 nm filamen DNA

DNA CHROMOSOME

Kromosom Sblm pembelahan sel (stlh sintesis DNA), kromatin mengalami pembelahan kondensasi mbtk individu kromosom metafase, yg nampak sbg sepasang kromatid (tampak spt 2 pasang lengan)

Kromosom Jmlh kromosom di dlm sel berbeda2 sesuai jns organismenya Btk & ukuran kromosom selama mitosis berubah2 , sebag bsr memp 2 lengan & tdpt lekukan dsbt kinetokor / sentromer Sentromer ini merup tempat melekatnya kromosom pd mikrotubul dr gelendong mitosis & sekaligus sbg pusat pergerakan kromosom pd stadium anafase. Kromosom tanpa sentromer dsbt kromosom asentrik & umumnya gagal memisahkan diri selama pembelahan sel. Kromosom saat interfase seakan2 hilang, yg tampak dlm nukleus hanya anyaman filamen halus, dsbt kromatin

TTAGGG Telomeres Gene 1 Gene 2 Telomeres DNA Chromosome TELOMERES in human beings are a repeating sequence of three of these nucleotide units: Thymine (T), Adenine (A), and Guanine (G), in the sequence: Important in replication - they act like a buffer zone Each time a normal cell divides its telomeres are cut shorter Cells with critically short telomeres become unable to divide further, and eventually malfunction and die Basically a chromosome is one long chain of genetic material. It is the microscopic, rod-shaped, threadlike part of the cell that carries hereditary information in the form of genes.  Every cell has chromosomes and all individuals in the same species have the same number of chromosomes.  Within an individual, every cell has the same number of chromosomes and generally come in pairs (except in sex cells).   Humans, for instance, have 23 pairs, or 46 chromosomes. A gene can be defined as a region of DNA that controls a hereditary characteristic. It usually corresponds to a sequence used in the production of a specific protein or RNA. Genes can be as short as 1000 base pairs or as long as several hundred thousand base pairs. It can even be carried by more than one chromosome. The estimate for the number of genes in humans has decreased as our knowledge has increased. As of 2001, humans are thought to have between 30,000 and 40,000 genes. telomere is a repeating DNA sequence (TTAGGG) at the end of the body's chromosomes. The telomere can reach a length of 15,000 base pairs. Telomeres function by preventing chromosomes from losing base pair sequences at their ends. They also stop chromosomes from fusing to each other. However, each time a cell divides, some of the telomere is lost (usually 25-200 base pairs per division). When the telomere becomes too short, the chromosome reaches a "critical length" and can no longer replicate. This means that a cell becomes "old" and dies by a process called apoptosis. Telomere activity is controlled by two mechanisms: erosion and addition. Erosion, as mentioned, occurs each time a cell divides. Addition is determined by the activity of telomerase. apoptosis: The process by which a cell dies at a natural, "pre-programmed" time Telomeres Chromosome

Telomere Length Declines 1,500 3,000 8,000 Age (years) 35 Telomere length in base pairs (human white blood cells) Telomere length declines in dividing cells as we age. In human blood cells, the length of telomeres ranges from 8,000 base pairs at birth to 3,000 base pairs as people age and as low as 1,500 in elderly people. (An entire chromosome has about 150 million base pairs.) Each time a cell divides, an average person loses 30 to 200 base pairs from the ends of that cell’s telomeres. Cells normally can divide only about 50 to 70 times, with telomeres getting progressively shorter until the cells become senescent, die or sustain genetic damage that can cause cancer. Telomeres do not shorten with age in tissues such as heart muscle in which cells do not continually divide. 65

A HISTORY OF DNA SEE p. 292-293 Discovery of the DNA double helix A. Frederick Griffith – Discovers that a factor in diseased bacteria can transform harmless bacteria into deadly bacteria (1928) B. Rosalind Franklin - X-ray photo of DNA. (1952) C. Watson and Crick - described the DNA molecule from Franklin’s X-ray. (1953)

DNA Double Helix “Rungs of ladder” Nitrogenous Base (A,T,G or C) “Legs of ladder” Phosphate & Sugar Backbone

UNIT FUNGSIONAL DNA/RNA

Perbedaan struktur gula antara RNA dan DNA

Basa DNA/RNA

IKATAN RANTAI DNA

IKATAN FOSFODIESTER

RNA is a polymer of ribonucleotides pGCUA C G U A 5’ end 3’ end 1’ 2’ 3’ 4’ 5’ OH PO-CH2 O H2N-C HN N CH O P O CH2 NH2 HC O-H RNA is a polymer of ribonucleotides ribose sugars Phosphodiester linkages Directional chain (5’ to 3’) 4 Bases purines: adenine & guanine pyrimidines: cytosine & uracil

DNA/RNA BERSIFAT ANTIPARALEL

DNA vs. RNA RNA DNA Rantai tunggal Double Helix Gula ribosa Uracil replaces Thymine! Ditransport ke sitoplasma DNA Double Helix Gula deoksiribosa Adenine pairs with Thymine (A-T) Tetap ada di nukleus

DNA is a polymer of 2’-deoxyribonucleotides pGCTA 5’ end 3’ end C G T A OPO-CH2 O H2N-C HN N CH O P O CH2 NH2 HC OH CH3 1’ 2’ 3’ 4’ 5’ DNA is a polymer of 2’-deoxyribonucleotides 2’-deoxyribose sugars Phosphodiester linkages Directional chain (5’ to 3’) 4 Bases purines: adenine & guanine pyrimidines: cytosine & thymine

RNA is easily hydrolyzed under alkaline conditions . O P O-CH2 N P . O P O-CH2 O N OH O P OH . O P O-CH2 O N OH O P O CH2 O P O ... H2O O N OH O P O ... HOCH2 mixture of 2’- and 3’- monophosphate derivatives H + RNA shortened RNA The reaction proceeds through a 2’,3’-cyclic monophosphate intermediate. Enzymatic hydrolysis of RNA by RNase proceeds through a similar intermediate. Because DNA lacks the 2’-OH group, it is stable under alkaline conditions.

Why does DNA contain T rather than U? Cytosine deaminates non-enzymatically to form uracil. If this happens in DNA, it constitutes a mutation. A proof-reading system recognize the error, and replaces the U by C. N CH C O HN NH2 cytosine uracil H2O Deamination of cytosine is of little consequence in RNA, which is not the permanent repository of genetic information.

The phosphate groups of DNA and RNA are negatively charged 5’ HO-CH2 O N O P O CH2 O-PO32 + M A phosphodiester group has a pKa of about 1, and so will always be ionized and negatively charged under physiological conditions (pH ~7). Nucleic acids require counterions such as Mg2+, polyamines, histones or other proteins to balance this charge. + M + M 3’ + M

The sugars are always in the b-furanose (cyclic) form 3’ 1’ 5’ 4’ endo C-2’ exo The ring can adopt various puckered conformations in which C-2’ and C-3’ are in either exo or endo positions relative to the base and C-5’. CH2OH H-C-OH H-C=O ribose in its aldehyde form b-furanose (ring) form HO-CH2 OH O The sugars are always in the b-furanose (cyclic) form 5’ 3’ HO-CH2 O N OH O P O CH2 O-PO32

The nucleotide base can rotate with respect to the sugar syn-Adenosine NH2 C N HC CH O HOCH2 OH anti-Adenosine syn-Cytosine anti-Cytosine The nucleotide base can rotate with respect to the sugar The bases can adopt either syn or anti conformations, but anti conformations are preferred.

3 TIPE DNA

BENTUK DNA

The B-form DNA helix has a diameter of about 20 Å ~20 Å Base pairs fill the center of the helix; the phosphates ( ) are on the outside. A base pair is more exposed to the solvent on one side (the “major groove”, at the top in these views) than the other (the “minor groove”, bottom).

B-form DNA consists of a right-handed double helix with antiparallel strands 5’ 3’ major groove minor groove minor 3.4 Å per bp 34 Å (10 bp) per turn 3’ 5’ These dimensions are for DNA fibers. In solution, there are ~10.5 base-pairs per turn.

Melting and Renaturation of DNA Renaturation driven by homologous base pairing

Untai ganda DNA A260 = 1.00 Untai tunggal DNA A260 = 1.37 Basa bebas A260 = 1.60

The two strands of the double helix separate reversibly at high temperatures The temperature at which this “denaturation” or “melting” occurs depends on the pH and salt concentration, and increases with the GC content of the DNA. (The curves drawn here are schematic.) If the temperature is lowered, the strands recombine. The rate of reassociation is inversely proportional to the complexity of the DNA.

Double-stranded and single-stranded DNA differ in their optical absorption at 260 nm dC dG dU dsDNA ssDNA nucleotides The conjugated p-electron systems of the purine & pyrimidine bases absorb strongly in the UV. (That’s why UV light is mutagenic and carcinogenic.) The absorbance of double-stranded DNA (dsDNA) at 260 nm is less than that of either single-stranded DNA (ssDNA) or the free bases. This is called “hypochromism.”

TERIMA KASIH