MBM Block 2

↑ rate of rxn w/o being consumed

↓ Ea

Do NOT change rxn equilibrium

Enzymes

ΔG < 0

Spontaneous

↑ [S] = ↑ rxn velocity until enzyme becomes saturated

x-axis = [S]

y-axis = rxn rate (V)

Michaelis-Menten

Maximum rxn velocity

Enzyme is saturated w/substrate

Vmax

[S] at ½ Vmax

Measure of enzyme affinity for substrate

Km

↑ Km =

LOW affinity

x-axis: 1/[S]

y-axis: 1/V

Lineweaver-Burk Plot

Oxidation-reduction

Add or remove electrons

Ex. dehydrogenase

Oxidoreductase

Transfer functional groups

Ex. kinase

Transferase

Cleave bonds w/ water

Ex. protease

Hydrolase

Cleave or add groups w/o hydrolysis or oxidation

Ex. decarboxylase

Lyase

Rearrange within a molecule

Ex. mutase

Isomerase

Joins two molecules w/ ATP

Ex. synthase

Ligase

Directly competes w/ substrate for enzyme active site

• Increasing substrate can overcome inhibition

↑ Km: need more substrate to achieve same velocity

Vmax: unchanged

Competitive Inhibition

Binds at allosteric site

↓ enzyme activity

↓ Vmax

Km: unchanged

• Increasing substrate cannot overcome inhibition

Noncompetitive Inhibition

Irreversibly inhibits enzyme

Ex. allopurinol

Suicide Inhibition

pH on Enzyme Activity

Optimal pH = HIGH activity

Extreme pH can cause denaturation

Temperature on Enzyme Activity

↑ T = ↑ Molecular Movement = ↑ Collisions = ↑ Rxn Rate

HIGH T = Denaturation = ↓ Enzyme Activity

Substrate Concentration of Enzyme Activity

LOW [S] = ↑ substrate = ↑ rxn rate

↑ activity does NOT equal ↑ rxn velocity

Inhibitors on Enzyme Activity

↓ Enzyme Activity

Reversible = inhibitor can dissociate

Irreversible: permanetly inactivates enzyme

Protein component only

Inactive

Apoenzyme

Non-protein component required for enzyme activity

Metal Ions: Mg2+, Zn2+, Fe2+/Fe3+, Cu2+

Cofactor

Organic non-protein molecule that assists an enzyme

Ex. NAD+, FAD+, CoA, TPP

Coenzyme

Coenzyme permanently associated w/enzyme

Ex. heme group of cytochromes, biotin group of acetyl-CoA carboxylase

Prosthetic Group

Apoenzyme + required nonprotein component

Active

Holoenzyme

Irreversible enzyme inhibitor → inhibits bacterial cell-wall synthesis → bacteria can’t maintain/build cell wall

Penicillin

Competitive inhibitors → resemble PABA → interfere with bacterial folate synthesis

Sulfonamides

Competitive inhibitor of folate metabolism → ↓ DNA synthesis/cell division → used as an anticancer drug

Methotrexate

Inhibits cytochrome oxidase

Cyanide

Inhibits enolase

Fluoride

↑ ALT =

Hepatocellular/parenchymal liver injury

↑ ALP =

Obstruction/cholestasis

↑ CK-MM =

Muscle Disease

↑ ALP with increased osteoblastic activity

Bone ALP is heat labile

Bone Disease

↑ PSA =

Prostate Disease/Cancer Marker

↑ Acid Phosphatase =

Prostate Cancer

Which enzyme rises first after a myocardial infarction?

CK-MB

Which enzyme rises after CK-MB in mycardio infarctions?

AST

Which enzyme rises last after a myocardial infarction?

LDH

LDH pattern with myocardial infarction

• Flipped LDH pattern

LDH1 > LDH2

Biomarker for myocardial infarction

Cardiac tropinin

CK-MB levels after myocardial infarction

Peak at 24 hrs

Baseline at 2-3 days

Gout =

↑ uric acid → urate crystal deposition → inflammation

Allopurinol

inhibits XO → ↓ uric acid → treats gout

Hemoglobin Structure

4 heme + 4 globin groups

Heme Synthesis Pathway

• δ-ALA Dehydratase Defiency

• Accumulation of porphyrin precursor ALA within RBCs and hepatocytes

• Triggers: alcohol, prolonged fasting

• Labs: lactic acidosis, elevated liver levels

• Symptoms: neurologic/GI failure

δ-ALA Dehydratase Porphyria

• PBG Deaminase Deficiency

• Ineffective heme synthesis + accumulation of PBG in cytoplasm

• Triggers: drugs

• Labs: elevated urine PBG + prophyrins

• Symptoms: neuropathic abdominal pain, DARK RED URINE, polyneuropathy, psychological disturbances

• Treatment: glucose + hematin to inhibit ALA synthase

Acute Intermediate Prophyria

• Uroporphyrinogen decarboxylase deficiency

• Triggers: hepatic iron overload, alcohol, sunlight, estrogen therapy, HBV/HCV

• Painful, blistering skin lesions on sun-exposed skin

• Onset during 4th/5th decade of life

• Red to brown urine

Porphyria Cutanea Tarda

• Protophorphyngogen oxidase deficiency

• In adulthood

• Triggers: nongenetic factors - drugs, diet, hormones, stress

• Symptoms: abdominal issues, muscle weakness, seizures, mental changes

• Photosensitivity

• Treatment: liver transplant

Variegate Porphyria

• In erythropoietic cells

• Enzyme deficiency in RBC

• Accumulation of protoporphyrins in RBC

• Symptoms: photosensitivity and liver disease

• In early childhood

Erythropoietic Protoporphyria

• Childhood Symptoms: developmental delays, abdominal issues, irritability

• Adult Symptoms: porphyria symptoms, anemia

• Interferes with ALA dehydratase and ferrochelatase → decreases heme synthesis

Lead Poisoning

Degradation of Heme

Heme → Biliverden → Bilirubin → Liver via Bilirubin-Albumin Complex → Bilirubin in Liver → Bilirubin Diglucuronide → to blood → Bile

↑ Serum bilirubin levels

Jaundice

Excess RBC breakdown/hemolysis

Prehepatic Jaundice

Problem w/ liver processing/conjugation

Hepatic Jaundice

Obstruction of bile flow

Posthepatic Jaundice

RBC Synthesis > RBC Destruction

Polycythemia

RBC Synthesis < RBC Destruction

Anemia

a globin chains are made on…

Chromosome 16

β globin chains are made on…

Chromosome 11

Hemoglobic Chain Levels

• Before Birth =

• After Birth =

• Before Birth = high γ chains, low β chains

• After Birth = low γ chains, high β chains

Sickle Cell Disease

Missense mutation in β-globin gene (Glu → Val)

HbS/HbS (homozygous)

Sickle Cell Trait

HbA/HbS (heterozygous)

No full clinical manifestation

Why do sickle cell symptoms occur after infancy?

Babies have high HbF

↓ HbF = ↑ β-globin production → ↑ HbS

Sickling Cause =

Low O2 + Dehydration → HbS polymerization

Sickling Hemolysis

Sickled RBCs are fragile = destroyed more easily

RBC desctruction → heme breakdown → ↑ bilirubin → jaundice

Sickling Vaso-occlusion

Sickles RBC → occlusion → tissue hypoxia → pain + tissue damage

Chronic hemolytic anemia

Lifelong episodes of pain

↑ infection sisceptibility

Acute chest syndrome

Stroke

Splenic dysfunction

Renal dysfunction

Bone marrow hyperplasia

Sickle Cell Disease Clinical Manifestations

1/4 defective genes

No physical manifestations

a-thalassemia silent carrier

2/4 defective genes

Mild anemia

a-thalassemia trait

3/4 defective genes

Hemolytic anemia

a-thalassemia HbH disease

4/4 defective genes

Fetal death

a-thalassemia hydrops fetalis

1/2 defective genes

Asymptomatic

β-thalassemia minor

2/2 defective genes

Severe disease and deformation

β-thalassemia Cooley Anemia

↑ pH → _____ shift → _____ affinity

LEFT shift → ↑ affinity

↓ temp, ↓ 2.3-BPG

↓ pH → _____ shift → _____ O2 → _____ affinity

RIGHT shift → Hb releases more O2 → ↓ affinity

↑ temp, ↑ 2,3-BPG

Deoxyhemoglobin

Hb w/o O2

Oxyhemoglobin

Hb w/ O2

Methemoglobin

Fe3+ instead of Fe2+

Carboxyhemoglobin

Hb bound to CO

Carbaminohemoglobin

CO2 bound to globin

Sulfhemoglobin

Abnormal Hb associated w/ certain sulfur-containing drugs

Glycohemoglobin

Glucose nonenzymativally attached to Hb

Glucosylation of hemoglobic occurs in…

Diabetes mellitus

Adjacent to cell membrane

Treadmilling: continuously adding at barbed end and dissociation at pointed end

Actin

Actin binding proteins (ABP)

Initiate formation, growth, stabilization

Formin, profilin, Arp2/3, WASP

Binds ATP-actin

Initiates nucleation

Adds actin to barbed end

Forms unbranches chains

Formin

Exchange ADP for ATP

Mutations linked to ALS

Profilin

Forms branches chains

Drives cell movement

Binds at barbed end

Arp2/3

Activated by Rho signaling pathway

Stimulates Arp2/3

Promotes polymerization @ leading edge

WASP

Cell shape, locomotion, intracellular transport, separation of chromosomes during mitosis

Microtubules Function

Microtubules Structure

• a- and β-tubulin dimers

• 13 protofilaments form hollow microtubule

• + and - end

• GTP hydrolysis: growth and shrinkage

Kinesin

Move toward + end

Dynein

Move toward (-) end

Generates sliding forces responsible for movement

Cilia Arrangement

9 outer doublets + 2 central microtubules

Provide mechanical strength and structural stability

Help cells withstand physical stress

More stable

Intermediate filament functions

Keratin are in…

Epithelial cells

Vimentin are in…

Mesenchymal cells

Desmin are in…

Muscle