Clostridial glutamate dehydrogenase mutants, designed to accommodate
the 2¢-phosphate of disfavoured NADPH, showed the expected large speci-ficity shifts with NAD(P)H. Puzzlingly, similar assays with oxidized cofac-tors initially revealed little improvement with NADP
, although rates with
were markedly diminished.
Đây là bài giảng của thầy Đỗ Hiếu Liêm, trường ĐH Nông Lâm TP.HCM. Chuỗi hô hấp mô bào bao gồm các phản ứng oxid hoá
khử liên tục trên các cơ chất (sản phẫm biến dưỡngtrung gian - Chu trình Krebs và β oxid hoá acid béo). Tổ chức: hệthống enzyme CHH với các coenzyme tổchức thành từng tổ hợp và sắp xếp theo thứ tự tăng dầnđiện thế oxid hoá khử,bắđầu NAD+ dehydrogenase có điện thế âm và cuối cùng là cytochrome oxidase có điện thế dương....
Early metabolic steps, including glycolysis and the activity of the pyruvate dehydrogenase complex, yield a two-carbon fragment called an acetyl group, which is linked to a large cofactor known as coenzyme A (or CoA). It is during the citric acid cycle that acetyl-CoA is oxidized to the waste product, carbon dioxide, along with the reduction of the cofactors NAD+ and ubiquinone
Glutamate dehydrogenase (EC 18.104.22.168–4) fromPeptostreptococcus asacch-arolyticushas a strong preference for NADH over NADPH as a coenzyme,
over 1000-fold in terms ofkcat
⁄Kmvalues. Sequence alignments across the
wider family of NAD(P)-dependent dehydrogenases might suggest that this
preference is mainly due to a negatively charged glutamate at position 243
(E243) in the adenine ribose-binding pocket.
Elucidating the origin of substrate and coenzyme specificity has been the
focus of much work relating to enzyme engineering. Many enzymes exhibit
tight specificity for particular substrates despite a close structural relation-ship to other nonreactive compounds.
The ubiquinone coenzyme Q (CoQ) is synthesized in mitochondria with a
large, hydrophobic isoprenoid side chain. It functions in mitochondrial res-piration as well as protecting membranes from oxidative damage. Yeast
that cannot synthesize CoQ (DCoQ) are viable, but cannot grow on nonfer-mentable carbon sources, unless supplied with ubiquinone.
Formate dehydrogenase from Candida boidinii(CboFDH) catalyses the
oxidation of formate anion to carbon dioxide with concomitant reduction
to NADH. CboFDH is highly specific to NAD
fails to catalyze the reaction with NADP
. Based on structural informa-tion for CboFDH, the loop region betweenb-sheet 7 and a-helix 10 in the
dinucleotide-binding fold was predicted as a principal determinant of coen-zyme specificity.
The biosynthesis of lovastatin inAspergillus terreusrequires two mega-synthases. The lovastatin nonaketide synthase, LovB, synthesizes the inter-mediate dihydromonacolin L using nine malonyl-coenzyme A molecules, and
is a reducing, iterative type I polyketide synthase.
The di-iron flavoprotein F420H2 oxidase found in methanogenic Archaea
catalyzes the four-electron reduction of O2to 2H2O with 2 mol of reduced
coenzyme F420(7,8-dimethyl-8-hydroxy-5-deazariboflavin). We report here
on crystal structures of the homotetrameric F420H2 oxidase fromMethan-othermobacter marburgensis at resolutions of 2.25 A
˚ , 2.25 A
˚ and 1.7 A
respectively, from which an active reduced state, an inactive oxidized state
and an active oxidized state could be extracted.
The functional activities of proteins are closely related to their molecular
structure and understanding their structure–function relationships remains
one of the intriguing problems of molecular biology. We investigated struc-tural changes in 17b-hydroxysteroid dehydrogenase from the fungus
Cochliobolus lunatus(17b-HSDcl) induced by pH, temperature, salt, urea,
guanidine hydrochloride, and coenzyme NADPH binding.
The roles of His181, His184 and Tyr186 in PETN reductase have been
examined by mutagenesis, spectroscopic and stopped-flow kinetics, and by
determination of crystallographic structures for the Y186F PETN reductase
and reduced wild-type enzyme—progesterone complex. Residues His181
and His184 are important in the binding of coenzyme, steroids, nitro-aromatic ligands and the substrate 2-cyclohexen-1-one.
The effect of CoA on the characteristic light decay of the firefly luciferase
catalysed bioluminescence reaction was studied. At least part of the light
decay is due to the luciferase catalysed formation of dehydroluciferyl-adenylate (L-AMP), a by-product that results from oxidation of luciferyl-adenylate (LH2-AMP), and is a powerful inhibitor of the bioluminescence
The Baeyer–Villiger monooxygenase (BVMO), 4-hydroxy-acetophenone monooxygenase (HAPMO), uses NADPH
to oxidize a variety of aromatic ketones and sulfides.
The FAD-containing enzyme has a 700-fold preference for
NADPH over NADH. Sequence alignment with other
BVMOs, which are all known to be selective for NADPH,
revealed three conservedbasic residues,whichcouldaccount
for the observed coenzyme specificity. The corresponding
residues in HAPMO (Arg339, Lys439 and Arg440) were
mutated and the properties of the purified mutant enzymes
were studied. ...