分子生物图谱

1、会计学1分子生物图谱分子生物图谱Prokaryotic Cell (does not have nucleus)Archaebacteria and eubacteria*Archea: Methnogens (anaerobic) Extreme halophiles (dead sea) Extreme thermophiles (hot spring, geysers)Eukaryotic Cell (contain a nucleus)Protista, fungi, animals and plantsCells:Cytoplasm: proteins, ribosome, meta

2、bolites and ionsPlasma membrane: phospholipid bilyer, associated proteins and carbohydratesDNAs, mtDNA: Gram-negative- Cell wall (3 layers: Periplasmic space; peptidoglycan; outer membrane) Gram-positive-Cell wall (thicker peptidoglycan layer) *Quite sensitive to lysozyme and penicillin The hydrophi

3、lic gel surrounding the cell wall in most bacteria Long, rigid protein roads, facilitating the movement of motile bacteria Short hair-like structure and attach other cells (essential infecting other organisms) A small, often unicelluar, reproductive unit of plants, algae, fungi, protoza, and bacteri

4、a Endoplasmic reticulumNuclear envelope NucleolusNucleoplasmLong arm: q (for queue)Short arm: p (for petit)Dark band: G band or G + band (AT rich)Light band: R band or G band (GC rich)(The membrane-bound structure in a cell)MitochondriaChloroplastsEndoplasmic reticulumGolgi apparatusPeroxisomesLysos

5、omesVacuolesGlyoxisomesSize: 1.5-2.0 in length, 0.5-1.0 um in diameterApprox the same as E. coliMaternal inheritanceMany copies: occupying of the cytoplasmic volumeRole: produce ATPEncode: proteins and RNAsmtDNAThylakoloids 类囊体Chlorophylls: located on the Thylakoloid membrane to absorb light for pho

6、tosynthesis Light 2H2O-O2 + 4H + +4e-H+ +ADP 3- + Pi 2- -ATP 4- + H2ORough ER: process newly synthesized peptides from ribosomeSmooth ER: involved in the synthesis and metabolism of lipids Major site for sorting and modifications of proteins and lipidsrough ER-transport vesicles-Golgi-GlycoproteinsP

7、eroxisome: contain enzymes for degrading amino acids and fatty aidsCatalase2H202-2H20 +02Lysosomes : 1) nuclease for degrading DNA and RNA 2) Protease for degrading proteins and other enzymes for degrading polysacchrides and lipids 3) lysosomes exist only in animal cells (plant -vacuoles for degradi

8、ng macromolecules) Found mainly in plant seedsFatty acids-Acetyl CoAMicrobodies: Peroxisomes +GlyoxisomesBaltimore ClassificationStructureLife CyclesThe life cycle of Viruses1 ) Attachment: viral surface protein-host cell receptor e.g: HIV GP120 - CD4 & Chemokine receptor (T cell)2) Penetration:

9、 endocytosis3) Uncoating: viral capsid degraded by viral enzyme or host enzyme4) Replication: assembly viral proteins and DNA or RNAs5) Release: escape from host cell by causing cell rupture (lysis) are viruses infect bacteriadsDNA phages with contractile tails, such as T4dsDNA phages with long flex

10、ible tails, such as dsDNA phages with stubby tails, such as P22ssDNA phages, such as Phi X174ssRNA phages, such as MS2 phages replicate rapidly and eventually cause lysis of the host cell the viral DNA circularizes and integrates into the host DNABacterium: Bacillus anthracisCutaneous (skin)Gastroin

11、testinalInhalationGram +2 major virulence factors: Poly-D-glutamic acid capsule: protecting from being killed by phagocyteAnthrax toxins: lethal factor edema factor protective antigen-cleaves members of mitogen-activated protein kinase kinase (MAPKK), interrupting the signaling pathways -adenylate c

12、yclase-mediates two factors by binding to a cellular receptor Building Blocksamino acidsPeptidesSecondary structureThree dimensional structureEnzymesMembrane proteinMiscellaneous proteinsAcidic: aspartic acid; glutamatic acidsBasic: lysine, arginine & histidineAromatic: tyrosine, tryptophan and

13、phenylalanineSulfur: cysteine, methionineUncharged hydrophilic: serine, threonine, asparagine, glutamineInactive hydrophobic: glycine, alanine, valine, leucine, isoleucineSpecial structure: prolineSalt bridge: interaction between + & - R groups, important stabilizing force in proteinsDisufide bo

14、nds: strong force for stabilizing the globular structureMethionine: synthesis of all peptide chains starts from methionineProline: the only amino acid has its R group and amino group directly connected- often located at the turn of a peptide in the 3-D structure of a protein is a chain of amino acid

15、s linked together by peptide bonds. usually refer to long peptides whereas are short peptides ( pIpH pIProtein structureCharge 1: Ion-exchange chromatographyColumn + anions+Sample mixtureProtein bindingColumn + proteinsColumn + anionsIon displacingPurified proteinProtein structure+Charge 2: Electrop

16、horesisProtein migrate at different position depending on their net chargeProtein structureCharge 3: Isoelectric focusingA protein will stop moving at position corresponding to its isoelectric point (pI) in a pH gradient gel. Protein structure3. Hydrophobicity: hydrophobic interaction chromatography

17、Similar to ion-exchange chromatography except that column material contains aromatic or aliphatic alkyl groupsProtein structure4. Affinity chromatographyEnzyme-substrate bindingReceptor-ligand bindingAntibody-antigen bindingProtein structure5. Recombinant techniques:Clone the interested protein enco

18、ding gene in an expression vector with a purification tag added at the 5- or 3 end of the gene Protein overexpression in a cell Protein purification with affinity chromatography.Protein structureMass Determination Gel filtration chromatography and SDS-PAGEComparing of the unknown protein with a prop

19、er standardPopular SDS-PAGE: cheap and easy with a 5-10% errorSDS: sodium dodecyl sulfate, makes the proteins negatively charged and the overall charge of a protein is dependent on its mass.Protein structureMass spectrometry:Molecules are vaporized and ionized, and the degree of deflection (mass-dep

20、endent) of the ions in an electromagnetic field is measuredextremely accurate, but expensiveMALDI can measure the mass of proteins smaller than 100 KDaHelpful to detect post-translational modificationProtein sequencing: relying on the protein data base Protein sequencingProtein sequencing : : Determ

21、ine the primary Determine the primary structure of a proteinstructure of a proteinSpecific enzyme/chemical cleavage: Trypsin cleaves after lysine (K) or arginine (R)V8 proteease cleaves after glutamic acid (E) Cyanogen bromide cleaves after methionine (M)Edaman degradation:Performed in an automated

22、protein sequencerDetermine the sequence of a polypeptide from N-terminal amino acid one by one.Expensive and laboriousProtein structureMost protein sequences are deduced from the DNA/cDNA sequence Direct sequencing: determine the N-terminal sequences or some limited internal sequence construction of

23、 an oligonulceotide or antibody probe fishing the gene or cDNAProtein structureX-ray crystallographyX-ray crystallography and and NMRNMRDeterming the tertiary structure (3-D) of a proteinDeterming the tertiary structure (3-D) of a proteinX-ray crystallography:X-ray crystallography: Measuring the pat

24、tern of diffraction of a beam of X-rays as it pass through a crystal. The first hand data obtained is electron density map, the crystal structure is then deduced. A very powerful tool in understanding protein tertiary structureMany proteins have been crystallized and analyzedProtein structureMeasuri

25、ng the relaxation of protons after they have been excited by radio frequencies in a strong magnetic field Measure protein structure in liquid but not in crystal Protein measured can not be larger than 30 KDaNMRNMRNucleotide: pentose +base + phosphateNucleoside: pentose +base A, C, G, T exist in DNAA

26、, C, G, U exist in RNA1 Overview of Gene Expression2 Overview of Transcription3 Genes Regulatory Elements4 Transcription Mechanism in Prokaryotes5 Transcription Mechanism in Eukaryotes 6 Motif Structure of Transcription Factors7 Histone Acetylation and DNA methylation 8 Regulation of Transcription Factors9 Transcription of RNA Genes10 Genome Replication of VirusesantisenseExon skippingEnzyme are the catalysts of biochemical reaction in the cellsNumerous drugs are enzyme inhibitors:HI