Biochemistry 3/e - Garrett & Grisham

Hecht Lecture 4:00 PM, Monday, April 2, 2012 Room 304 Chemistry Building

Can DNA Function as a Natural Molecular Switch in Cells?

Professor Laurence Hurley University of Arizona
This is a must see It will be a substitute for our regularly scheduled lecture on Tues April 3 You will be responsible for handling questions on this presentation material on the exam If you cannot make the seminar, get the notes from someone who did
2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Chapter 24
Lipid Biosynthesis
to accompany Biochemistry, 3/e by Reginald Garrett and Charles Grisham

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Outline
24.1 Fatty Acid Biosynthesis 24.2 Biosynthesis of Complex Lipids - touch on 24.3 Eicosanoid Biosynthesis - touch on 24.4 Cholesterol Biosynthesis 24.5 Transport via Lipoprotein Complexes

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Overview of Lipid Biosynthesis

Large portion of the metabolic map Of immense importance with a variety of compounds that include:
fatty acids sphingolipids eicosanoids cholesterol steroids

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Fatty Acid Pathways

- The Biosynthesis and Degradation Pathways are Different -

As seen with glycolysis/gluconeogenesis and with glycogen synthesis/breakdown ---fatty acid synthesis and degradation go by different routes There are four major differences between fatty acid breakdown and biosynthesis
Oxidation -SH Intermediates Enzymes Energy Source
Linked to CoA Separate enzymes Uses NADH/NAD+

Synthesis
Linked to acyl carrier prPart of one polypeptide Uses NADPH/NADP+

Location
2005 Brooks/Cole - Thomson

Mitochondria

Cytosol

Biochemistry 3/e - Garrett & Grisham

Activation by Malonyl-CoA
Fatty acids are built from 2-C units
acetyl-CoA

Acetate units are activated for transfer by conversion to malonyl-CoA

costs ATP

Decarboxylation of malonyl-CoA and reducing power of NADPH drive chain growth Chain grows to 16-carbons Other enzymes add double bonds and/or more Cs

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Overall reaction

The Enzymes of Lipid Biosynthesis

Shuttling
citrate synthase malate DH PDH pyr carboxylase ATP-citrate lyase
Malonyl-CoA

transient carbonyl-phosphate int.

carboxybiotin

Synthesis - two components

acetyl-CoA carboxylase fatty acid synthase complex
malonyl transferase -ketoacyl-ACP reductase dehydratase

Activate bicarb

Acetyl-CoA carbanion

Biotindependent carboxylation
Nucleophilic attack

Malonyl-CoA
2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Acetyl-CoA Carboxylase
- "ACC enzyme" commits acetate to fatty acid synthesis ACC the only FA biosynth. component NOT part of the enzyme complex Carboxylation of acetyl-CoA to form malonyl-CoA is the irreversible, committed step in FA biosynthesis SO WHAT? regulated! ACC uses bicarbonate, ATP and biotin Animal enzyme is one polypeptide with all three functions - biotin carboxyl carrier, biotin carboxylase and transcarboxylase (E.coli enzyme has three separate subunits) read how regulated.

Acyl-CoA

Activated bicarb

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

FA Synthesis
- Separate enzymes in a complex in Bacteria/Plants Not a multi-enzyme complex Decarboxylation drives condens. rxn of Ac-CoA and mal-CoA Other three steps
reduction-dehydration-reduction Familiar?? exact reverse of FA degradation (see pg 747)

transfer Create ACP conjug.

B-ketoacylACP Synthase
transfer Create ACP conjug.

reduction

Domain 1

dehydration

Domain 2

reduction

Two major differences

D (not L) configuration NADPH

Net cost for synthesis of Palmitoyl-CoA (see equations on page 770) Difference vs -oxidation?
7 ATP 14 NADPH where from?

2005 Brooks/Cole - Thomson

Product

Biochemistry 3/e - Garrett & Grisham

Fatty Acid Synthesis in Animals

- Fatty Acid Synthase (FAS) - a multienzyme complex -

Dimer of identical 250 kD subunits each subunit a multifunctional polypeptide - arranged in head-to-tail fashion Three functionally separate domains Domain 1
acetyl transferase malonyl transferase -ketoacyl ACP synthase

Domain 2
dehydratase -ketoacyl reductase enoyl-ACP reductase

Domain 3
thioesterase
2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Fatty Acid Synthesis in Animals

- Fatty Acid Synthase (FAS) - a multienzyme complex -

Dimer of identical 250 kD subunits each subunit a multifunctional polypeptide - arranged in head-to-tail fashion Three functionally separate domains Domain 1
acetyl transferase malonyl transferase -ketoacyl ACP synthase

Domain 2
dehydratase -ketoacyl reductase enoyl-ACP reductase

Domain 3
thioesterase
2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Fatty Acid Synthesis in Animals

- Fatty Acid Synthase (FAS) - a multienzyme complex -

Note the roles of active site serines on AT & MT Note the roles of ACP and KSase Steps 3-6 repeat to elongate the chain

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Regulation of FA Synthesis
- Allosteric modifiers Malonyl-CoA inhibits carnitine acyltransferase
blocking uptake of fatty acylcarnitine by mitochondria and, thus, inhibits -oxidation

Citrate activates acetyl-CoA carboxylase Fatty acyl-CoAs inhibit acetyl-CoA carboxylase

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Regulation of FA Synthesis
- Phosphorylation & Hormones Hormones regulate ACC thru post-translational modification Glucagon Glucagon
activates lipases inhibits ACC
associated with low blood glucose

active

Insulin
inhbits lipases activates ACC
SYNTHESIS
RECIPROCAL inhibited

BREAKDOWN

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Biosynthesis of Other Lipids

Phosphatidic acid is the precursor for all other glycerolipids in eukaryotes
PA is made either into DAG or CDP-DAG precursor for synthesis of:
PI, PC, PE, PS, cardiolipin (pg. 783)

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Phosphatidic Acid

http://www.topnews.in/health/files/depression4
2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Biosynthesis of Other Lipids

Cardiolipin
present in metabolically active cells of the heart and skeletal muscle a "double" phospholipid
it has four fatty acid tails

serves as an insulator and stabilizes the activity of protein complexes important to the ETS

bisphosphatidyl glycerol

a "double" phospholipid
it has four fatty acid tails

Clinical Significance
Barth syndrome
rare genetic disorder recognised in 1970's to cause infantile death a mutation in the gene coding for the enzyme synthesizing cardiolipin sufferers have mitochondria that are abnormal, and cannot sustain adequate production of ATP

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Biosynthesis of Other Lipids

Cardiolipin
present in metabolically active cells of the heart and skeletal muscle a "double" phospholipid
it has four fatty acid tails

serves as an insulator and stabilizes the activity of protein complexes important to the ETS

bisphosphatidyl glycerol

a "double" phospholipid
it has four fatty acid tails

Clinical Significance
Barth syndrome
rare genetic disorder recognised in 1970's to cause infantile death a mutation in the gene coding for the enzyme synthesizing cardiolipin sufferers have mitochondria that are abnormal, and cannot sustain adequate production of ATP
http://www.lipidprofiles.com/images/nrdd_2005_fig01.gif

2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Biosynthesis of Other Lipids

Ceramide a key precursor for complex lipids
sphingolipids
if glycosylated get cerebrosides
cerebrosides + sialic acid get gangliosides

Function
component of neural tissues, membrane stabilizer immune recognition (ABO), toxin recognition (cholera)

Glycosphingolipids
Sphingomyelin Lecithin/Sphingo (L/S) ratio Abnormal in Tay-Sachs disease
2005 Brooks/Cole - Thomson

Biochemistry 3/e - Garrett & Grisham

Biosynthesis of Other Lipids

Ceramide a key precursor for complex lipids
sphingolipids
if glycosylated get cerebrosides lecithin
cerebrosides + sialic acid get gangliosides

Function
component of neural tissues, membrane stabilizer immune recognition (ABO), toxin recognition (cholera) Mature

L:S Ratio

PG

>1.9
1.5-1.9 <1.5

Present
Absent Absent

Glycosphingolipids

Sphingomyelin Lecithin/Sphingo (L/S) ratio Immature Mature Abnormal in Tay-Sachs (diabetic mothers) disease
2005 Brooks/Cole - Thomson

Borderline

>3.4