Biochemistry

Biochemistry
Processes
- Blood clotting
- Vitamin K
- Function
- Add carboxylic acid group to a
glutamate amino acid residue
- form a gamma-carboxyglutamate
(Gla) residue
- Usage
- VKOR reduces to vit K hydroquinone
- K hydroquinone oxidised
- Allows carboxylation of Glu to Gla
- By gamma-glutamyl carboxylase
- Also vit-K dependens carboxylase
- Only happens if oxidation to vit
K epoxide works
- Linked reaction
- Vitamin K epoxide is then
reconverted to vitamin K by
VKOR
- Reduction and subsequent reoxidation of vitamin
K coupled with carboxylation of Glu is called the
vitamin K cycle
- Enzyme function
- Features
- Protein/RNA catalysts
- Specificity
- Regulation
- Enzyme expression
- Enhancers
- Inhibitors
- Catalytic power
- Lower activation energy
- Additional components
- Cofactors
- Coenzymes
- If tightly bound
- = Prosthetic group
- Group bound = holoenzyme
- Group unbound = apoenzyme
- Calculations
- v = d[A]/dt = K[A]
- A+B > P+Q
- v= dA/dt = dB / dt = dP /dt = dQ / dt
- v = k[A][B]
- ES <k1 / k-1 > E+S
- (k-1)[ES]<>(k1)[E][S]
- Ks=[ES]/[E][S] = (k-1)/(k1)
- E+S <k1/k-1> ES >k2> E+P
- Km = (K1+K2)/(K-1)
- v =
- Substrate binding
- H bond
- Ionic
- Vd Waals
- Inhibitors
- Effects
- Km
- Vmax
- Reversible
- Competitive
- Same binding site as substrate
- Km up
- Noncompetitive
- Pure
- IE and E binding to S are same
- Km same
- Vmax down
- Mixed
- IE-S binding altered
- Close to binding site
- Conformational change
- Interact with E and ES
- Uncompetitive
- I binds ES only
- Km up
- Vmax down
- Irreversible
- Suicide substrate
- Permanent binding (covalent)
- Glycogen
- Glycogenolysis
- Glycogen(n residues) + Pi <>
glycogen(n-1 residues) + glucose-1-phosphate
- 1. Glycogen phosphatase
phosphorilates a-1-4
- Until 4 residues before a-1-6 branch
- Glygogen debranching puts 3
onto another branch
- α[1→6] glucosidase hydrolises
last glucose off
- Only case of non G1P
- Hexokinate converts it to G1P
- 2. G1P to G6P
- by phosphoglucomutase
- Glycogenesis
- 1. Glucose to G6P
- Glucokinase
- Hexokinase
- 2. G6P to G1P
- Phosphoglucomutase
- glucose-1,6-bisphosphate`intermediate
- 3. G1P to UDP-Glucose
- Uridyl Transferase
- aka UDP-glucose pyrophosphorylase
- Pyrophosphate formed
- Hydrolyses to 2 Pi
- Pyrophosphatase
- 4. Chain assembly
- Glycogen synthase
- Start on old chain
- Or glycogenin (primer)
- Process
- Binds to UDPG
- Oxonium ion forms
- Add to 4 hydroxyl group
- 5. Branching
- amylo-α(1:4)→α(1:6)transglycosylase
- End of chain onto earlier part
pH
- Hyperventilation
- Increased gass/lung contact
- Lower CO2
- Lower H2CO3
- Lower H+ + HCO3-
- Alkidosis
- Vasoconstriction
- Hb affinity for O2 increases
- Lower O2
- Formulas
- pH = -Log[H+]
- [H+] = 10^-pH
- [H+] = ([HA]Ka)/[A-]
- [H+][OH-]=10^-14 M
- Weak acid
- Ka = [H+][A-] / [HA]
- Ka low = weak acid
- pKa = -log(Ka)
- pKa+pKb=14
- pH=pKa + log([A-]/[HA])
- [A-] = X
- [HA] = Concentration-X
Specific proteins
- Hemoglobin
- Hemoglobin saturation curve
- CO2 decreases O2 affinity
- BPG lowers O2 affinity
- First O2 molecule binds
- heme Fe ion is drawn into the
porphyrin ring plane
- Confirmational change to subunits
- Enhance O2 affinity
- H+ promotes dissociation
- Deoxyhemoglobin
- Higher H+ affinity
- Some CO2 transported directly
- O NH-COO
- Fetal blood has lower BPG affinity
- Hogher O2 affinity
- Structure
- 2 a-units
- 2 b-units
- Each unit has heme group with Fe
- Trouble
- Sicle cell disease
- Advangates
- Protection against malaria
- Characteristics
- Hemoglobin mutation
- b-Globulin chain mutation
- Glutamic acid is replaced
- Hydrophobic valine
- Confirmational change
- @Low oxygen
- Non-covalent polymerisation
- Myoglobin
- Behavior
- Binds 1 O2
- Traditional Michaelis-menten graph
- Oxygen storage in muscle
- Collagen
- Structure
- Connective tissue
- Prolyl hydroxylase
- Needs vit C to function
- Cofactor to keep Fe++ in reduced state
- Involved in collagen synthesis
Protein structure
- Levels
- Primary
- Aminoacid sequence
- Aminoacids
- Only L configuration in bio
- 20 aminoacids in human code
- Amino group
- -NH2
- Acid group
- -COOH
- Peptide bond
- -C(=O)NH-
- Secondary
- Alpha helix
- Right hand coiled
- Backbone NH H links to C=O
- 4 residues earlier
- 3.6 resudues per turn
- Beta sheet
- Lateral connection
- At least 2-3 bonds per line
- Configurations
- Parallel
- Anti-parallel
- No covalent bonds
- Tertiary
- 3D structure
- Non-specific hydrophoibic interactions
- Salt bridges
- H bonds
- Disulfide bonds
- Tight packing of side chains
- Quarternaru
- Multi tertiary units
- Same noncovalent bonds as tertiary
- Multiple polypeptides
- Multimere
- Di/trimer etc
- Problems
- Prions
- Transmissable confirmational changes
- PrP
- PrP(c) is ok
- Membrane
- a-Helical
- Cell adhesion & communication
- PrP(sc) is not ok
- Mutated
- Higher proportion of beta sheets
- Clogs up cell
- Death
Membrane transport
- Passive diffusion
- Active transport
- Transporter proteins
- Primary
- Pump
- ATP
- Secondary
- Uniport
- Cotransport
- Symport
- Antiport
- Gradient
Genetics
- Gene expression
- Transcription
- Phases
- Pre-initiation
- Initiation
- Promoter clearence
- Elongation
- Gyrase introduces negative
super coils
- Reduce strain
- Topoisomerase to remove
negative coilks after bubble
- Termination
- Rho factor (enzyme)
- Termination site (sequence)
- Steps
- DNA to mRNA
- Recognition of promoter (TATAAT)
- Sigma factor
- RNA-polymerase binds
- Helicase enzymes make
transcription bubble
- RNA polymerase runs
- 3'UTR poly-A tail and a 5'UTR cap
- Translation
- Initiation
- 30S ribosome complex binds
- IF3-30S-IF1
- IF-2 delivers tRNA-fMet into P site
- GDP bound
- 50S subunit binds
- GTP hydrolysed
- IF-2 dissociated
- Results in 70S subunit
- Elongation
- Complex shit, will add if bored
- Termination
- Stop codon found
- RF-/RF2 +RF-3:GTP binds to A site
- Confirmational jumbo
- Dissociation
- RNA
- Can cleave own backbone
- 2' hydroxyl oxygen attacks the
phosphodiester bond
- Uracil
- Replaces thymine in RNA
- Uracil methylation results in Thymine
- Deaminated cytosine is uracil
- Uracil DNA glycosilate fixes
- Evolved before DNA
- Types
- Mainstream
- mRNA
- tRNA
- rRNA
- ncRNA
- Lesser known
- miRNA
- siRNA
- piRNA
- snRNA
- snoRNA
- dsRNA
- DNA
- PCR
- Step 1
- Isolate target sequence
- Heat to 95 celcius
- Cool to 70
- Step 2
- Add 1000 fold excess primer
- Step 3
- (Taq) DNA polymerase
- Heat resistant
- dTTP, dATP, dGTP, dCTP
- Step 4
- Heat to 95
- Cool to 70
- Step 5
- Repeat 4
- Forensic PCR
- DNA profiling
- VNTRs
- variable number tandem repeats
- STR
- particularly short tandem repeats
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