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1、Obligatory course for biology studentsPrerequisite: General biology, Biochemistry Molecular BiologyProkaryotic Cell (does not have nucleus)Archaebacteria and eubacteria*Archea: Methnogens (anaerobic) Extreme halophiles (dead sea) Extreme thermophiles (hot spring, geysers)Eukaryotic Cell (contain a n
2、ucleus)Protista, fungi, animals and plantsCells:Cytoplasm: proteins, ribosome, metabolites and ionsPlasma membrane: phospholipid bilyer, associated proteins and carbohydratesDNAs, mtDNABasic Cellular ComponentsBasic Cellular ComponentsProkaryotic cells Prokaryotic cells BacteriaBacteria: Gram-negati
3、ve- Cell wall (3 layers: Periplasmic space; peptidoglycan; outer membrane) Gram-positive-Cell wall (thicker peptidoglycan layer) *Quite sensitive to lysozyme and penicillinCapsule and slime:Capsule and slime: The hydrophilic gel surrounding the cell wall in most bacteriaFlagella:Flagella: Long, rigi
4、d protein roads, facilitating the movement of motile bacteriaFimbriaeFimbriae and and pilipili: : Short hair-like structure and attach other cells (essential infecting other organisms)Spores:Spores: A small, often unicelluar, reproductive unit of plants, algae, fungi, protoza, and bacteria Organizat
5、ion of Eukaryotic CellsOrganization of Eukaryotic CellsEndoplasmic reticulumBiomembraneBiomembraneNucleusNucleusNuclear envelope NucleolusNucleoplasmChromosome and Chromosome and karyotypekaryotypeLong arm: q (for queue)Short arm: p (for petit)GimsaGimsa ( (QuinacrineQuinacrine) )Dark band: G band o
6、r G + band (AT rich)Light band: R band or G band (GC rich)Organelles in the CytoplasmOrganelles in the Cytoplasm(The membrane-bound structure in a cell)MitochondriaChloroplastsEndoplasmic reticulumGolgi apparatusPeroxisomesLysosomesVacuolesGlyoxisomesMitochondriaMitochondriaSize: 1.5-2.0 in length,
7、0.5-1.0 um in diameterApprox the same as E. coliMaternal inheritanceMany copies: occupying of the cytoplasmic volumeRole: produce ATPEncode: proteins and RNAsmtDNAChloroplastsChloroplastsThylakoloids 类囊体Chlorophylls: located on the Thylakoloid membrane to absorb light for photosynthesis Light 2H2O-O
8、2 + 4H + +4e-H+ +ADP 3- + Pi 2- -ATP 4- + H2OEndoplasmic reticulumEndoplasmic reticulumRough ER: process newly synthesized peptides from ribosomeSmooth ER: involved in the synthesis and metabolism of lipids GolgiGolgi apparatus apparatus Major site for sorting and modifications of proteins and lipid
9、srough ER-transport vesicles-Golgi-GlycoproteinsPeroxisomesPeroxisomesPeroxisome: contain enzymes for degrading amino acids and fatty aidsCatalase2H202-2H20 +02LysosomesLysosomesLysosomes : 1) nuclease for degrading DNA and RNA 2) Protease for degrading proteins and other enzymes for degrading polys
10、acchrides and lipids 3) lysosomes exist only in animal cells (plant -vacuoles for degrading macromolecules) GlyoxisomesGlyoxisomes 乙醛酸循环体乙醛酸循环体Found mainly in plant seedsFatty acids-Acetyl CoAMicrobodies: Peroxisomes +GlyoxisomesVirusesVirusesBaltimore ClassificationStructureLife CyclesThe life cycl
11、e of Viruses1 ) Attachment: viral surface protein-host cell receptor e.g: HIV GP120 - CD4 & Chemokine receptor (T cell)2)Penetration: 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
12、causing cell rupture (lysis) BacteriophagesBacteriophagesBacteriophagesBacteriophages are viruses infect bacteriadsDNA phages with contractile tails, such as T4dsDNA phages with long flexible tails, such as dsDNA phages with stubby tails, such as P22ssDNA phages, such as Phi X174ssRNA phages, such a
13、s MS2LyticLytic cycle: cycle: phages replicate rapidly and eventually cause lysis of the host cell LysogenicLysogenic cycle: cycle: the viral DNA circularizes and integrates into the host DNALife cycleLife cycleAnthrax and Biological WeaponsAnthrax and Biological WeaponsBacterium: Bacillus anthracis
14、Cutaneous (skin)GastrointestinalInhalationPathogenesis of AnthraxPathogenesis of AnthraxGram +2 major virulence factors: Poly-D-glutamic acid capsule: protecting from being killed by phagocyteAnthrax toxins: lethal factor edema factor protective antigenLethal factorLethal factor-cleaves members of m
15、itogen-activated protein kinase kinase (MAPKK), interrupting the signaling pathways Edema factorEdema factor-adenylate cyclaseProtective antigenProtective antigen-mediates two factors by binding to a cellular receptor Protein structure and functionProtein structure and functionBuilding Blocksamino a
16、cidsPeptidesSecondary structureThree dimensional structureEnzymesMembrane proteinMiscellaneous proteinsThe 20 amino acids of proteinsThe 20 amino acids of proteinsAcidic: aspartic acid; glutamatic acidsBasic: lysine, arginine & histidineAromatic: tyrosine, tryptophan and phenylalanineSulfur: cystein
17、e, methionineUncharged hydrophilic: serine, threonine, asparagine, glutamineInactive hydrophobic: glycine, alanine, valine, leucine, isoleucineSpecial structure: prolineSalt bridge: interaction between + & - R groups, important stabilizing force in proteinsDisufide bonds: strong force for stabilizin
18、g 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 proteinPeptidePeptide is a chain of amino acids linked together
19、by peptide bonds.PolypeptidesPolypeptides usually refer to long peptides whereas oligopeptidesoligopeptides are short peptides ( pIpH pIProtein structureCharge 1: Ion-exchange chromatographyColumn + anions+Sample mixtureProtein bindingColumn + proteinsColumn + anionsIon displacingPurified proteinPro
20、tein structure+Charge 2: ElectrophoresisProtein 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: hydro
21、phobic interaction chromatographySimilar 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:
22、Clone the interested protein encoding 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 o
23、f the unknown protein with a proper 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 t
24、he degree of deflection (mass-dependent) 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 sequen
25、cingProtein sequencing : : Determine 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 degra
26、dation:Performed in an automated 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
27、internal sequence construction of an oligonulceotide or antibody probe fishing the gene or cDNAProtein structureX-ray crystallographyX-ray crystallography and and NMRNMRDetermingDeterming the tertiary structure (3-D) of a protein the tertiary structure (3-D) of a proteinX-ray crystallography:X-ray c
28、rystallography: Measuring the pattern 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
29、 analyzedProtein structureMeasuring 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: pento
30、se +base BaseBaseA, C, G, T exist in DNAA, C, G, U exist in RNAThe nucleic acid chainThe nucleic acid chainDNA 3D structureDNA 3D structureDNA 3D structureDNA 3D structureRNA StructureRNA StructureGene TranscriptionGene Transcription1 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 skipping
