Milk Biosynthesis PART 2: ENERGY Key Enzymes (from all biosynthesis lectures) FDPase = fructose diphosphatase

Citrate lyase

Isocitrate dehydrogenase

Fatty acid synthetase

Acetyl CoA carboxylase

Fatty acyl deacylase – thioesterase II

Lipoprotein lipase NAD, NADH NADH ◦ Nicotinamide adenine dinucleotide ◦ Reduced form ◦ Pyridine nucleotide that functions as an oxidative cofactor in eukaryotic cells ◦ Plays a key role in the production of energy through redox reactions NAD ◦ Oxidized form ◦ Serves as a cofactor for dehydrogenases, reductases, and hydroxylases ◦ Major carrier of H+ and e- in major metabolic pathways like glycolysis, the TCA cycle, and fatty acid synthesis NADH important in glycolysis and Citric Acid cycle NADPH is used in cytosol for fatty acid synthesis

Energy and Carbon Sources

Mammary Whole C-Source Blood C-Source Gland Body Lactose Glucose Fatty Acids Primary Primary Source Energy Energy Source Source Ruminant Glucose Acetate Glucose Propionate Acetate and butyrate Monogastric Glucose Glucose Glucose Diet Glucose

Glycolysis Embden Meyerhof Pathway Metabolic pathway used to begin breakdown to glucose Purpose: TO PRODUCE PYRUVATE AND ATP

Pentose Phosphate Pathway Alternative pathway of glucose metabolism

Generates pentoses, particularly ribose for use in nucleic acid synthesis

Produces NADPH

◦ Tissues requiring large amounts of NADPH produced by this pathway are the tissues that synthesize fatty acids and steroids (e.g. mammary gland, adipose tissue, adrenal cortex and liver)

◦ Tissues less active in fatty acid synthesis such as skeletal muscle are virtually lacking the pentose phosphate pathway

Works in two phases: oxidative and non-oxidative

Pentose Phosphate Pathway: Oxidative

Glucose-6-phosphate oxidized into ribulose-5-phosphate (and CO2) During oxidation of glucose-6-phosphate, NADP+ is reduced to NADPH The second step of the pathway coverts the ribulose 5-phosphate into other pentose-5- phosphates including ribose-5-phosphate used to synthesize nucleic acids The third step converts three of the pentose-5-phosphates into two molecules of hexoses and one triose In the fourth step, some of these sugars are converted into glucose-6- phosphate so the cycle can be repeated ◦ Three-carbon sugar is not recycled in the monogastric without FDPase (reverse glycolysis) Pentose Phosphate Pathway Summary

Oxidative phase: 3Glucose-6-phosphate + 6NADP+ →2 Xylulose-5p + + ribose-5P + 3CO2 + 6NADPH + 6H

Rearrangements of the nonoxidative phase: 2 Xylulose-5P + ribose-5P → 2 Frucose-6P + Glyceraldehyde-3P

Sum of these two phases: 3 Glucose-6-phosphate + 6NADP+ → 2 + Frucose-6P + Glyceraldehyde-3P + 3CO2 + 6NADPH + 6H Pentose Phosphate Pathway Summary: Ruminants vs Monogastrics Monogastrics: 3Glucose-6-phosphate + 6NADP+ → 2 Frucose-6P + Glyceraldehyde-3P + + 3CO2 + 6NADPH + 6H Triose-P goes to TCA cycle Ruminants:

12 glucose-6-P + 12 NADP -> 6 CO2 + 5 6-carbon units recycled + 12 NADPH More glucose goes in but you recycle more carbons (due to FDPase) and still only use up 6 carbons released as CO2 Presence of FDPase allows recycling of triose-P (glyceraldehyde-3-P) so you produce 12 NADPH after the complete oxidation of one glucose equivalent (6 carbons) Ruminants are more efficient at producing NADPH (for fatty acid synthesis) than monogastrics

Citric Acid Cycle AKA Krebs cycle or tricarboxylic acid cycle (TCA cycle)

Purpose: TO PRODUCE ENERGY!

Starting point = pyruvate from Embden-Meyerhof pathway

Produces 30 ATP for each molecule of glucose oxidized

For each NADPH2 or NADH2 produced, 3 ATP are generated

Aerobic process

C3H4O3 + ½ O2 + 15 ADP + 15 Pi -> 3 CO2 + 2 H2O + 15 ATP Citric Acid Cycle Videos https://www.youtube.com/watch?v=F6vQKrRjQcQ https://www.youtube.com/watch?v=Fcu_8URp4Ac

FDPase

Citrate lyase (8) Hey!! Don’t forget about me! I’m used for energy production, too!! Energy and Carbon Sources

Mammary Whole C-Source Blood C-Source Gland Body Lactose Glucose Fatty Acids Primary Primary Source Energy Energy Source Source Ruminant Glucose Acetate Glucose Propionate Acetate and butyrate Monogastric Glucose Glucose Glucose Diet Glucose

What’s the Main Purpose? Glucose can go one of two ways:

Glycolysis (Embden Meyerhof Pathway): To produce pyruvate (2 pyruvates for every glucose) which will enter Citric Acid Cycle

Pentose Phosphate: To produce NADPH for fatty acid synthesis and ribose sugars for nucleic acid synthesis

Pyruvate goes to:

Citric Acid Cycle: To produce energy (NADH and FADH will eventually make ATP)

◦ Also produces oxaloacetate, citrate, isocitrate, malate, and 2-oxoglutarate (we will see these in the fatty acid lesson!) FDPase

Citrate lyase (8) Questions?