Chapter 25 -Biosynthesis of Amino Acids

Microorganisms use ATP and a powerful reductant to reduce atmospheric nitrogen to _______

ammonia

Nitrogen in biomolecules comes from ____ _____,N2.

atmospheric nitrogen

N2 has an extremely strong ____ bond that is highly resistant to chemical attack

triple

Nitrogen fixation

a process that reduces N2 to NH3 (ammonia)

_______ (nitrogen-fixing) microorganisms fix ~60% of N2

diazotrophic

Lightning and ______ radiation fix 15% of N2

ultraviolet

______ processes fix 25% of N2

industrial

Biological nitrogen fixation is catalyzed by the _____ _____

nitrogenase complex

Nitrogenase complex

a complex enzyme with multiple redox centers that fixes N2 into NH3

Nitrogenase complex requires ___ _____

ATP hydrolysis

The nitrogenase complex consists of two components:

– _______ (iron protein, Fe protein) = provides electrons with high reducing power

– ________ (molybdenum-iron protein, MoFe protein) = uses electrons to reduce N2 into NH3

reductase; nitrogenase

_____ ______ which supplies ATP for nitrogen fixation, requires O2.

oxidative phosphorylation

The nitrogenase complex is extremely sensitive to _______ by O2.

inactivation

Nitrogenase is located in the root nodules of ______ plants

leguminous

Leguminous plants maintain a very low concentration of free O2 in their root nodules, using an O2-binding protein that is a hemoglobin homolog (leghemoglobin).

– allows simultaneous functioning of ATP synthesis and ______

O2; nitrogenase

Reductase and nitrogenase are ___-____ proteins

iron–sulfur

The iron–molybdenum _____ of nitrogenase binds and reduces atmospheric nitrogen

cofactor

_______ transfers electrons from reduced ferredoxin to nitrogenase

– The ___-____ cluster of reductase carries electrons one at a time.

– ATP binding and hydrolysis trigger a confirmational change that moves reductase closer to ______.

reductase; 4Fe-4S; nitrogenase

Electrons from reductase enter at P clusters and flow to the ____ cofactor, the site of nitrogen fixation

FeMo

The Fe protein, or reductase, transfers electrons of high _____ power

reducing

Ammonium ion is assimilated into an amino acid through _____ and ______

glutamate; glutamine

NH3 generated by the ______ ______ becomes NH4+ in aqueous solutions

nitrogenase complex

Glu and Gln act as _____ donors for most amino acids.

– Glu contributes its α-amino group by ______ in the synthesis of most amino acids.

– Gln contributes its side-chain nitrogen atom in the synthesis of __ and __

nitrogen; transamination; Trp; His

Glutamate dehydrogenase

catalyzes the synthesis of glutamate from NH4+ and α-ketoglutarate

Glutamate dehydrogenase requires either NADH or NADPH and proceeds through a ____ _____ intermediate that becomes protonated and is subsequently reduced

NADH; NADPH; Schiff base

Glutamate dehydrogenase produces glutamate in two steps

• A Schiff base forms between ______ ion and α- ketoglutarate.

• The protonated Schiff base is reduced by the transfer of a _____ ion from NAD(P)H, forming glutamate

ammonium; hydride

Glutamate ______ is established by hydride transfer to the Schiff base

chirality

Glutamate dehydrogenase binds the _____ substrate such that the hydride transferred from NAD(P)H is added to form the __ isomer of glutamate

α-ketoglutarate; L

Glutamine synthetase incorporates another nitrogen into glutamate in an amidation reaction, forming _____

– requires ____ hydrolysis

– The reaction proceeds through an _______ intermediate

glutamine; ATP; acylphosphate

Catalyzes the reductive amination of α-ketoglutarate to glutamate and uses glutamine as the nitrogen donor

Glutamate synthase

When _____ is limiting, most of the _______ is made by the sequential action of glutamine synthetase and glutamate synthase

NH4+; glutamate

In nitrogen fixation:

a. electrons flow from ferredoxin to nitrogenase to the reductase.

b. The Fe protein transfers electrons of low reducing power.

c. ATP hydrolysis within the nitrogenase drives conformational changes necessary for electron transfer.

d. P clusters store electrons until they can be used to reduce nitrogen at the FeMo cofactor.

e. N2 is reduced to NH4+.

d

Amino acids are made from _______ of the citric acid cycle and other major pathways

intermediates

The majority of amino acids other than ___ and ___ obtain their nitrogen from Glu or Gln

Glu; Gln

______ _____ for amino acid synthesis are provided by intermediates of glycolysis, the citric acid cycle, or the pentose phosphate pathway

carbon skeletons

Which of the major metabolic precursors of amino acids are from the pentose phosphate pathway?

a. phosphoenolpyruvate

b. ribose 5-phosphate

c. erythrose 4-phosphate

d. α-ketoglutarate

e. 3-phosphoglycerate

b and c

Essential amino acids

9 amino acids that humans cannot synthesize and must be supplied in the diet

Nonessential amino acids

11 amino acids that humans can synthesize if dietary content is insufficient

____ is sometimes designated as essential, but it can be synthesized from available Phe in one step

Tyr

Aspartate, alanine, and glutamate are formed by the addition of an amino group to an ____-_____

alpha-ketoacid

α-Ketoglutarate can be converted into glutamate by _____ ______

reductive amination

Aspartate and alanine can be made from the addition of an ____ ____ to oxaloacetate and pyruvate respectively

• These transamination reactions are catalyzed by _____ _____-dependent aminotransferases.

amino group; pyridoxal phosphate

Almost all aminotransferases that participate in amino acid biosynthesis are related to aspartate aminotransferase by ______ evolution

divergent

Conserved residues include:

– the ___ residue that forms the Schiff base with the PLP cofactor

– the ____ residue that interacts with the α-carboxylate group of the ketoacid to orient the substrate

Lys; Arg

A key step in the transaminase reaction is the protonation of the quinonoid intermediate to form the external _____

aldimine

Chirality of the amino acid formed is determined by the _____ from which this proton is added

direction

The L configuration forms at the Cα center when the proton is transferred to the bottom face of the _______

intermediate

Proton addition during transamination establishes amino acid _________

stereochemistry

The formation of asparagine from aspartate requires an ______ intermediate

adenylated

Asn forms from _____ in an amidation reaction that is driven by ATP hydrolysis

Asp

The nitrogen donor for Asn synthesis is ____ in bacteria and ____ in mammals

NH4+; Gln

________ is the precursor of glutamine, proline, and arginine

glutamate

Glutamate is converted into _____ γ-semialdehyde, a precursor for both proline and arginine.

– requires ATP and _____

glutamic; NADPH

__-_______ is the precursor of serine, cysteine, and glycine.

3-phosphoglycerate

______ is synthesized from the glycolytic intermediate 3- phosphoglycerate

serine

3-Phosphoglycerate is oxidized to 3-phosphohydroxypyruvate, ________ to 3- phosphoserine, and hydrolyzed to serine

transaminated

Serine is the precursor of ______

glycine

Tetrahydrofolate

a highly versatile carrier of one-carbon units

In the formation of glycine, the side-chain methylene group of serine is transferred to ________

• catalyzed by the ____ enzyme serine hydroxymethyltransferase

tetrahydrofolate; PLP

Tetrahydrofolate is composed of 3 groups:

– a substituted _____ ring.

– p-aminobenzoate.

– a chain of one or more _______ residues

pteridine; glutamate

Mammals obtain ________ from their diets or from microorganisms in their intestinal tracts

tetrahydrofolate

The one-carbon group carried by tetrahydrofolate is bonded to its ___ or ____ nitrogen atom (denoted as N5 and N10) or to both

N-5; N-10

Tetrahydrofolate is an important carrier of _____ ___-____ ______

activated one-carbon units

One-Carbon Groups Carried by Tetrahydrofolate

Oxidation state:

• Most reduced (= ______)

• Intermediate (= ________)

• Most oxidized (= ____ ____)

methanol; formaldehyde; formic acid

N5,N10-methylenetetrahydrofolate donates a one-carbon unit in an alternative synthesis of _____ that starts with _____ and NH4+

• catalyzed by glycine synthase

• requires CO2, NH4+, and NADH

glycine; CO2

S-adenosylmethionine is the major donor of ____ groups

methyl

S-adenosylmethionine (SAM)

an activated methyl donor with higher transfer potential than tetrahydrofolate

S-adenosylmethionine (SAM) is synthesized from ______ and ______

methionine; ATP

S-adenosylmethionine is converted to _______

homocysteine

After donation of a methyl group by S-adenosylmethionine, the resulting S-adenosylhomocysteine is hydrolyzed, yielding ________ and homocysteine

adenosine

Methionine is regenerated by transfer of a methyl group to homocysteine from N5-methyltetrahydrofolate.

– catalyzed by _____ ______

– mediated by the coenzyme methylcobalamin, which is derived from vitamin ____

methyl; methionine synthase; B12

_______ is synthesized from serine and homocysteine

cysteine

Serine and homocysteine condense to form cystathionine, which is catalyzed by cystathionine β-synthase

cystathionine

Cystathionine is deaminated and cleaved to _____ and α-ketobutyrate

– catalyzed by cystathionine γ-____or cystathionase

cysteine; lyase

High homocysteine levels correlate with _____ disease

vascular

Elevated serum levels of ______ or the disulfide-linked dimer homocystine are a predisposing factor for coronary heart disease and arteriosclerosis

homocysteine

Elevated homocysteine levels commonly result from ______ in the gene encoding cystathionine β-synthase

mutations

Glyphosate

a broad-spectrum herbicide

Acts as an uncompetitive inhibitor of the enzyme that produces 5-enolpyruvylshikimate 3-phosphate and blocks aromatic amino acid biosynthesis in plants

glysophosphate

Chorismate is converted into ______, the immediate precursor to the aromatic ring of Phe and Tyr

• catalyzed by ______ mutase

prephenate; chorismite

Prephenate can be:

– dehydrated and decarboxylated to yield phenylpyruvate and transaminated to ___

– oxidatively decarboxylated to p-hydroxyphenylpyruvate and transaminated to ___

Phe; Tyr

Phenylalanine and tyrosine are formed in the _______ branch

prephenate

Chorismite acquires an amino group from Gln and releases ______, forming anthranilate

pyruvate

The carbon skeleton of _____replaces glyceraldehyde 3-phosphate in indole 3-glycerol phosphate, forming Trp.

– catalyzed by _____ synthase

serine; tryptophan

Tryptophan is formed in the _______ branch

anthranilate

Tryptophan synthase is an _____ tetramer

α2β2

The α subunit of tryptophan catalyzes the formation of ____ from indole-3-glycerol phosphate

indole

The β subunit of tryptophan has a _____-containing active site that catalyzes the condensation of indole and Ser to form Trp

PLP

Ser forms a Schiff base with PLP, which is dehydrated to yield a Schiff base of _______

aminoacrylate

Aminoacrylate is attacked by indole to yield ____

Trp

Tryptophan synthase uses substrate _______ to prevent loss of the indole intermediate

channeling

During the synthesis of most amino acids, a transamination reaction occurs. What property of amino acids is established following this reaction?

chirality

In feedback inhibition, the final product in a pathway inhibits the enzyme catalyzing the ______ step.

– conserves building blocks and metabolic energy

committed

The committed step in Ser synthesis is catalyzed by __-_________ dehydrogenase, which is inhibited by Ser

3-phosphoglycerate

The binding of Ser to a regulatory site of 3-phosphoglycerate reduces the value of ____ for the enzyme

Vmax

Threonine deaminase

the PLP enzyme that catalyzes the formation of α-ketobutyrate

Threonine deaminase is allosterically inhibited by ___ , the end product of its pathway and allosterically activated by ____, the end product of a competitive pathway

Ile; Val

Isozymes

enzymes with essentially identical catalytic mechanisms but different regulatory properties

Enzyme multiplicity

process in which the committed step can be catalyzed by two or more isozymes

Process by which a common step is partly inhibited by multiple final products, each acting independently

cumulative feedback inhibition