Biochem Ch 1, 3, 6

Biochem Ch 1, 3, 6

1. Chapter 1 AA, Peptides, Proteins

   • 1.1 Amino Acids Found in Proteins
     • A note on Terminology
       • Proteinogenic AA
         • 20 AA in genome
     • Stereochem of AA
       • All AA are L
         • Most AA are S
           • Cysteine is an exception
     • Structure
       • Non polar
         • Special
           • Methionine has sulfer atom but a Ch3 attached
             • Proline is cyclic with amino attached to the alpha
       • Aromatic: Tryptophan, phenylalanine, tyrosine
       • Polar
         • Special
         • Acid (-)
         • (+)
           • Histidine PH = 6
     • Phobic and Philic
   • 1.2 Acid-Base Chem of AA
     • Protonation & deprotonation
       • zwitterions: both + & - values
       • + in acid
       • - in base
     • Titration
       • PI: value where molecules are electrically neutral
         • PI= (PKA + PKA2)/2
     • Amphoteric: accept or donate a proton
   • 1.3 Peptide Bond Formation and Hydrolysis
     • di = 2 tri = 3 oligo <20 poly > 20
     • formation
       • condensation reaction puts AA together
         • AKA dehydration
       • breaks and uses water (hydrolysis)
     • hydrolysis
       • breaks, trypsin and chymotrypsin will help
   • 1.4 Primary and Secondary Protein Structure
     • beta make fibroin silk fibers
     • proline used for cell membrane crossing
   • 1.5 Tertiary and Quaternary Pro. Structure
     • R groups used for tertiary
       • 2 cysteine= cystine
     • Conjugated proteins
       • Use prosthetic groups
         • Lipoproteins, glycoproteins, nucleoproteins
   • 1.6 Denatureation
     • Disrupt by heat or solutes

2. Chapter 3 Nonenzymatic Protein Function and Protein Analysis

   • 3.1 Cellular functions
     • Structural
       • for cytoskeleton and helps support tissues such as ligaments.
       • Motif: super secondary creating a structure
       • Types
         • Collagen: For connective tissue
           • 3 left handed helices makeing a right handed helix
         • Elastin: part of the matrix, it stretches and recoils like a spring
         • Keratin: hair and nails,
         • Actin: microfilaments and thin in myofibrils., allows movement of motor proteins. (polar)
         • Tubulin: makes up microtubules. structure, separation, transport (polarity)
     • Motor
       • ATPases power change for proteins
       • Myosin interacts with actin, making it a part of cell transport.
       • Kinesin
         • Aligning chromosomes
         • Move transmitter to +
       • dyneins
         • move cilia and flagella
         • Move to -
           • waste or recycled neurotransmiter
     • binding proteins
       • Hemoglobin, and other affinities affect each one.
     • Cell adhesion Molecules
       • cadherins
         • run on Ca2+
         • hold similar cell types together
       • integrins
         • alpha and beta chains
         • Communicate with extracellular matrix
           • promote devision, apoptosis, platelet formation
       • selectins
         • weakest
         • bind to carbohydrates
           • sometimes on other cells
         • role: defense, inflammation
     • Immunoglobulins
       • 3 defenses
         • neutralize
         • opsonization: mark for destruction
         • Agglutinating
   • 3.2 Biosignaling
     • ion channels
       • facilitated = passive
       • ungated
       • Voltage gated (mem potential)
       • Ligand gated
         • gated open/closed due to binding
     • Enzymed linked
       • membrane spanning
       • ligand binding
         • activates domain/ conformational change
         • catalytic domain
     • GPCR
       • Heterotrimeric G: for transmitting signals
       • s= adenylate cyclase =increases cAMP
       • i= inhibits adenylate cyclase= decrease cAMP
       • q= activates phospholipase C, cleaves phospho out and into DAG. Open endoplasmic ret ca2+
   • 3.3 Protein Isolation
     • homogenization: crush grind, bleed.
     • electrophoresis
       • polyacrylamide gel
         • allow small to move faster
       • native PAGE
         • used for native states of protein.
       • SDS- PAGE
         • separates by mass
           • discrupts interactions and makes large - charges
       • Isoelectric focusing
         • focus on acid base properties
     • Chromatography
       • for large amounts of protein
       • material to stationary phase (adsorbent)
         • run --> elute
       • Column
         • break size and polarity
         • ion-exchange
           • charged beads will break charge flow
         • Suze-Exclusion
           • beads have small pores so large will move faster.
         • Affinity
           • receptors or bonds which hold compounds will slow or grab to desired material.
   • 3.4 Protein Analysis
     • Protein Structure
       • Edman degradation
         • breaks N terminal of 50-70 AA proteins
     • AA composition
     • activity Analysis
       • reacting colors and observing substrates help find activity
     • Concentration Determination
       • UV spec
       • BCA assay, lowry and bradford protein assay have colorimetric changes in reactions.
       • bradford: uses blue dye
         • protonated becomes brown-green
           • color change amount can give concentration

3. Chapter 6 DNA and Biotechnology

   • 6.1 DNA structure
     • Nucleosides and nucleotides
       • Nucleosides: 5 carbon pentose bonded to a base
       • Nucleotides: 1 or more phosphate is attached to C-5' side
       • deoxyribose missing oxygen at C-2
     • Shugar phos backbone
       • Read in 5-3' position
       • Nucleotides are joined at 3'-5'
     • Purines and Pyrimidines
       • Pu= 2 rings = A & G
         • Pure As Gold
       • Py = 1 ring = C & T & U
         • Cut The Py
     • Watson-Crick
       • DNA Helix
       • Chargaff's = G=C A=T
       • Most DNA is B
         • Z is also there, it is left handed and has high GC
     • Denatureation and Reannealing
       • Probe DNA is used to anneal and find evidence
   • 6.5 Recombinant DNA and Biotech
     • DNA cloning and Restriction Enzymes
       • Recombinant DNA tech allows for more gene cloing/ multiply
     • Safety and Ethics
     • Applications of DNA Technology
       • Gene Terapy = potential cures for ingerited diseases.
       • Transgenic and knockout mice
         • altered at germ line
           • transgene: is gene entered
         • knockout mice: intentionally deleted gene
         • could result in chimera
           • both new gene and old resulting in patches.
     • DNA Sequencing
       • dideoxyribonucleotide used to stop binding of polymerase
         • allows for fragments and size to be read.
     • Hybridization
       • Joining of complementary base pair sequences
       • PCR makes copies without amplifying bacteria DNA
         • 1. denature, 2 replicate 3 cool to reanneal 4 repeat
       • Gel electrophoresis and southern blotting
         • detect presence and quantitiy of DNA.
           • it will be probed to label.
         • use agarose gel. Longer = slower
     • DNA Libraries and cDNA
       • made my reverse transcribing mRNA
       • lack noncoding introns= expression library
   • 6.2 Eukaryotic Chromosome Organization
     • Histones
       • H2A, H2B, H3 H4 = Histone core
         • + 200 DNA = Nucleosome
       • H1 Seals DNA as it enters and leaves the nucleosome.
       • nucleoproteins
         • proteins that associate with DNA
     • Heterochromatic & Euchromatin
       • Heterochromatin Compacted DNA is dark
       • Euchromatin = light and active DNA
     • Telomeres and Centromeres
       • Telomere
         • TTAGGG
         • Reapeating unit which is used for loss of sequence.
       • Centromeres = center of chromosome (high GC)
   • 6.3 DNA Replication
     • Strand Separation and Origins of Replication
       • Helicase Unwinds DNA
       • SSDNA binding proteins unravel strand
       • Topoisomerases make negative supercoil but cutting ahead
     • Synthesis of Daughter Strands
     • Replicating the Ends of Chromosomes
       • telomers are used to allow cutting of DNA.
         • have high GC content
   • 6.4 DNA Repair
     • Oncogenes and Tumor Suppressor Genes
       • cancer moves by metastasis
       • Oncogenes are cancer causeing mutations
         • proto oncogen= before mutated
         • antioncogenes: repress tumors, issues if 2 repressor genes.
     • Proofreading and Mismatch Repair
       • Proofreading
         • Starts with polymerase enzyme
       • mismatch
         • in G2
           • by MSH2 and MLH1
             • similar to MutS & MutL
     • Nucleotide and Base Excision Repair
       • damage recognized in H1 and G2
       • UV light forms thimine dimers
         • NER used to cut and patch
           • 1. recognize 2. excision endonuclease makes nick in phosphodiester
             • 3. DNA polymerase will fill gap
               • 4. DNA ligase fix nicks
       • Base Excision
         • 1. recognized and removed by glycosylase
           • 2. AP site (abasic) recognized by AP endonuclease and removes damage from DNA
             • 3. DNA polymerase and ligase
   • Genomic libraries have all DNA in an organisms Genome
     • cDNA is only expressed genes

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