10.FA14 Metabolic Engineering

Metabolic BiochemistryBIBC102

Lecture 10

October 24, 2014

Metabolic flux and its control

1 Glucose (C6H12O6)

2 GAP

2 PYR

2 LACTIC ACID ( 2 x C3H6O3)

2 ADP + 2 Pi

2 ATP

Biological Work

Feedback Inhibition

LNC 15.16 (3rd ed.)

“bottleneck”)

We are not dealing with isolated reactions

at equilibrium, but with a flux of carbons

through a pathway

this flux must be regulated and adjusted to

physiological needs

near equilibrium

LNC 3rd ed.

LNC 15.18a

active site

active site

regulatory site

regulatory site

Phosphofructokinase 1

(only two of four subunits shown)

Fru 6-P + ATP Fru 1,6-bisphosphate + ADP

LNC 15.18b

ATP is both a substrate

anda negative modulator

LATERLecture 20

AMP stimulates the enzyme !!!!ATP inhibits the enzyme !!!!

High AMP

Low AMP

AMP concentrations

fluctuate more widely, and therefore the sensitivity

of the enzyme can be increased

if it responds to changing

AMP concentrations

The role of adenylate kinase:

2 ADP ATP + AMP Keq = 0.44

[ATP] [AMP]Keq = 0.44 =

[ADP]2

when [ATP] is high, [AMP] is low

when [ATP] is low, [AMP] is high

LNC 4th ed.

Typical physiological concentrations

A relatively small decrease in ATPcauses

a relatively large increase in AMP

A researcher (Catalina Island Marine Institute) scuba diving in the Toyon Bay found this dead fish that normally

lives 3000 feet below the surface.

We do not normally eat GLUCOSE,

but we eat carbohydrates/junk food

in various forms

Other Common Sugars

Monosaccharides: fructosegalactosemannose

Disaccharides: sucrose Glu-Frulactose Gal-Glumaltose Glu-Glutrehalose Glu-Glu

(Section 14.2) Feeder Pathways for Glycolysis)

Disaccharides

Figure 7-12

Galactose Glucose

Figure 7-12

GlucoseFructose

Figure 7-12

Glucose Glucose

Formation of an ETHER

R1 - CH2 - O - H + H - O - CH2 - R2

R1 - CH2 - O - CH2 - R2

H2O

Lactose + H2O D-galactose + D-glucose

Sucrose + H2O D-fructose + D-glucose

Trehalose + H2O 2 D-glucose

Lactase

Sucrase

Trehalase

Enzymes on surface of intestinal epithelial cells

Lactose IntoleranceOMIM

Online Mendelian Inheritance in Man

Major symptoms: diarrhea, dehydration, bloat

Remedy: Lactaid (lactase) taken orally

http://www.ncbi.nlm.nih.gov/omim?cmd=search

In humans, the activities of lactase and most of the other digestive hydrolases are maximal at birth. The majority of the world's human population experiences a decline in production of the digestive enzyme lactase-phlorizin hydrolase during maturation, with the age of onset ranging from the toddler years to young adulthood. Due to the reduced lactase level, lactose present in dairy products cannot be digested in the small intestine and instead is fermented by bacteria in the distal ileum and colon. The fermentative products result in symptoms of diarrhea, gas bloat, flatulence, and abdominal pain. However, in a minority of adults, high levels of lactase activity persist in adulthood. Lactase persistence is a heritable autosomal dominant condition that results in a sustained ability to digest the milk sugar lactose throughout adulthood (Olds and Sibley, 2003).

Clinical FeaturesMontgomery et al. (1991) stated that in those humans who have low lactase activity as adults, the decline occurs at approximately 3 to 5 years of age. They suggested that the developmental pattern of lactase expression is probably regulated at the level of gene transcription.

Adult-onset lactase decline appears to be a risk factor for developing osteoporosis, owing to avoidance of dairy products or interference of undigested lactose with calcium absorption (Lee and Krasinski, 1998). Elderly with both adult-onset lactase decline and atrophic gastritis or those undergoing anti-ulcer treatment may have an increased risk of low calcium absorption owing to the lack of gastric acid that facilitates calcium uptake.

Lactase and Human Evolution

• http://evolution.berkeley.edu/evolibrary/news/070401_lactose

The metabolism of galactose presentsanother problem

The solution, discovered by the Argentinian

scientist Leloir, included the discovery ofthe important class of compounds called

SUGAR NUCLEOTIDES

Lactose + H2O D-galactose + D-glucose

Galactose

1

Gal – O – P - O

Glu – O – P – O – P – O - RIBOSE - URACIL

Glu – O – P – O Glu 1-P

Gal – O – P – O – P – O - RIBOSE - URACIL

Gal 1-P

UDP-Galactose UDP-Glucose

epimerase

the reaction occurs in two steps (oxidation/reduction reactions)

oxidation

reduction

NADH + H+

NAD+

Figure 14-13

NADH + H+

NAD+

Mistake in Lehninger 4th edition and in e-book, but not in printed 5th and 6th edition

Mistake in Lehninger 4th edition, in e-book, in PowerPoint slides provided to instructors, but not in printed 6th edition

NAD+

NADH + H+

NADH + H+

NAD+

one single enzyme

no net oxidation/reduction

overall

Galactose Galactose 1-P

UDP-Glu

UDP-Gal

Glucose 1-P

Glucose 6-P

kinase

Uridyl transferase

epimerase

phosphoglucomutase

ATP ADP

Enzymes required for galactose metabolism

Glu 6-P ↔ Glu 1-P

Phosphoglucomutase

Galactosemiahigh galactose levels in blood

Possible enzyme deficiencies- galactokinase- transferase- epimerase- (mutase?)

Symptoms:large liver, icterus (jaundice), failure to thrive, and urinary excretion of

albumin and sugar; vomiting or diarrhea usually begins within a few days of milk ingestion

End of Lecture 10

October 24, 2014