Notes- Synthesis of Cycloheptatriene

Winberg, H. E.

doi:10.1021/jo01084a635

Cited by 46 publications

(17 citation statements)

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“…The structure of UDP-galactase-4-epimerase [40] shows NAD' oriented as reported for dihydropteridine reductase. UDP-4-galactose-epimerase is homologous to animal and viral 3&hydroxysteroid dehydrogenases, Nocardia cholesterol dehydrogenase, and plant dihydroflavonol reductases [4]. This expands the relevance of information about the residues important in catalysis by ADH.…”

Section: Discussionmentioning

confidence: 77%

“…In particular, Qr-153, Lys-157, and Thr-186 interact with either nicotinamide or its ribose. Tyr-153 is proposed to be the catalytically active residue [4,18,19]; Lys-157 is proposed to lower the pK, of tyrosine to facilitate catalysis at neutral pH [19]. Another role for Lys-157 has come from analysis of the tertiary structure of dihydropteridine reductase [24].…”

Section: Discussionmentioning

confidence: 99%

“…Oxidation of alcohols by ADH proceeds by an ordered reaction [3,4,37] in which the binding of NAD' induces a conformational change in ADH that promotes substrate binding and catalysis. Fig.…”

Section: Discussionmentioning

confidence: 99%

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Adding a positive charge at residue 46 of Drosophila alcohol dehydrogenase increases cofactor specificity for NADP⁺

et al. 1994

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We previously reported that the D39N mutant of Drosophila alcohol dehydrogenase (ADH), in which Asp-39 is replaced with asparagine, has a 60-fold increase in athnity for NADP' and a lJ-fold increase in k,, compared to wild-type ADH [Chen et al. (1991) Eur. J. B&hem. 202, 263-267) and proposed that this part of ADH is close to the r-phosphate on the ribose moiety of NADP+. Here we report the effect of replacing Ala-46 with an argine residue, an A46R mutant, on binding of NADP+ to ADH and its catalytic efficiency with the NADP' cofactor, and a modeling of the three-diiensional structure of the NAD'-binding region of ADH. The A46R mutant has a 2.5-fold lower Xm(lpp)nADP+ and a 3-fold higher /c_~ with NADP' compared to wild-type ADH, binding of NAD' to the mutant was unchanged and km, with NAD' was lowered by about 30%. For the A46R mutant, the ratio of k-,/G of NAD+ to NADP+ is 85, over ten-fold lower than that for wild-type ADH. Our model of the 3D structure of the NAD'-binding region of ADH shows that Ala46 is over 10 A from the ribose moiety of NAD+, which would suggest that there is little interaction between this residue and NAD+ and explain why its mutation to arginine has little effect on NAD+ binding. However. the positive charge at residue 46 can neutralize some of the coulombic repulsion between Asp-39 and the r-phosphate on the ribose moiety of NADP+, which would increase its a%nity for the A46R mutant. We also constructed a double mutant, D39N/A46R mutant, which we find has a 30-fold lower K&lppjN~Dp+ and 8-fold higher k,, with NADP' as a cofactor compared to wild-type ADH; binding of NAD+ to this double mutant was lowered by 5-fold and k_* was increased by 1.5-fold. As a result, k_&,, for the double mutant was the same for NAD+ and NADP+. The principle effect of the two mutations in ADH is to alter its at%ity for the nucleotide cofactor; k,, decreases slightly in A46R with NAD+ and remains unchanged or increases in the other mutants.Key words: Drosophila ADH; NADP' binding to ADH; Steroid and prostaglandin dehydrogenase IrltrodnctlonDrosophila alcohol dehydrogenase (ADH) has a preference for secondary alcohols [l-7] and does not require a metal ion for catalysis, in contrast to the yeast and horse liver alcohol dedhydrogenases. Drosophila ADH shares a common ancestor [8-121 with human 1 I&hydroxysteroid dehydrogenase, 17/Ihydroxysteroid dehydrogenase, and 15-hydroxyprostaglandin dehydrogenase; enzymes that regulate the concentration of glucocorticoids, estrogens, and prostaglandins, respectively, in humans and are thus of importance in various endocrine-related diseases [13-151.The mechanism of catalysis of ADH and its homologs is only beginning to be understood. The importance of the N-terminus of ADH in binding the nucleotide cofactor was first shown when Thatcher sequenced an inactive ADH mutant and found that Gly-15 was replaced by aspartic acid [16]. Site-specific mutagenesis studies with cloned Adh also indicate that the Nterminus is important in binding of the nucleotide cofactor [5,...

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Section: Discussionmentioning

confidence: 77%

Section: Discussionmentioning

confidence: 99%

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Adding a positive charge at residue 46 of Drosophila alcohol dehydrogenase increases cofactor specificity for NADP⁺

et al. 1994

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“…This reaction is analogous to the conversion of dichloronorcarane into cycloheptatriene (XXIII) (145).…”

Section: XXIImentioning

confidence: 99%

Carbenes. Reactive Intermediates Containing Divalent Carbon.

Chinoporos¹

1963

Chem. Rev.

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“…The yield in the formation of 7,7-dichlorobicyclo [4.1.01heptane varies from 59 to 79% depending on the source of dichlorocarbene. These bicyclic compounds (norcaranes) are of interest since on pyrolysis they yield cycloheptatrienes (25) CH, cx -0+ 6' 1 2 (57% total yield) Ring enlargement of alkenes in the heterocyclic series is also possible. For example dihydropyran may be converted into 2,3-dihydro-6-ehlorooxepine in 83% yield (based on the dichlorocyclopropane) (26) Benzocyclobutenone derivatives may be obtained from 1,4-cyclohexadienes produced by the Birch reduction of p-dimethoxybenzenes ( 27 The adduct formed on standing for 24 hr with silver sulfate and sulfuric acid gives a 35% yield of 3-bromo-4-hydroxybenzocyclobutenone.…”

Section: Cclfmentioning

confidence: 99%

Carbenes in insertion and addition reactions

Buehler¹

1972

J. Chem. Educ.

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Notes- Synthesis of Cycloheptatriene

Cited by 46 publications

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Adding a positive charge at residue 46 of Drosophila alcohol dehydrogenase increases cofactor specificity for NADP⁺

Adding a positive charge at residue 46 of Drosophila alcohol dehydrogenase increases cofactor specificity for NADP⁺

Carbenes. Reactive Intermediates Containing Divalent Carbon.

Carbenes in insertion and addition reactions

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Notes- Synthesis of Cycloheptatriene

Cited by 46 publications

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Adding a positive charge at residue 46 of Drosophila alcohol dehydrogenase increases cofactor specificity for NADP+

Adding a positive charge at residue 46 of Drosophila alcohol dehydrogenase increases cofactor specificity for NADP+

Carbenes. Reactive Intermediates Containing Divalent Carbon.

Carbenes in insertion and addition reactions

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Product

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Adding a positive charge at residue 46 of Drosophila alcohol dehydrogenase increases cofactor specificity for NADP⁺

Adding a positive charge at residue 46 of Drosophila alcohol dehydrogenase increases cofactor specificity for NADP⁺