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Literature summary extracted from

  • Li, S.; Ouellet, H.; Sherman, D.H.; Podust, L.M.
    Analysis of transient and catalytic desosamine-binding pockets in cytochrome P-450 PikC from Streptomyces venezuelae (2009), J. Biol. Chem., 284, 5723-5730.
    View publication on PubMedView publication on EuropePMC

Crystallization (Commentary)

EC Number Crystallization (Comment) Organism
1.14.15.33 structures of the PikC D50N mutant cocrystallized with narbomycin, to 1.85 A resolution, and with 10-deoxymethymycin, to 3.2 A resolution. In the mutant D50N, the desosamine moiety of both 10-deoxymethymycin and narbomycin is bound in a catalytically productive buried site, suggesting a two-step substrate binding mechanism, whereby desosamine is recognized in the two subsites to allow the macrolide substrate to sequentially progress toward a catalytically favorable orientation Streptomyces venezuelae

Protein Variants

EC Number Protein Variants Comment Organism
1.14.15.33 D50N increase in catalytic activity. Binding affinity for 10-desoxymethymycin increases about 4fold, binding affinity for narbomycin about 1.3fold compared to wild-type. 60% conversion of 10-desoxymethymycin, ratio of products neomethymycin to methymycin is 1.15. 44.4% conversion of substrate narbomycin Streptomyces venezuelae
1.14.15.33 D50N/E85Q 18.4% conversion of 10-desoxymethymycin, ratio of products neomethymycin to methymycin is 0.38. 3.5% conversion of substrate narbomycin Streptomyces venezuelae
1.14.15.33 D50N/E94Q no conversion of 10-desoxymethymycin. 5.3% conversion of substrate narbomycin Streptomyces venezuelae
1.14.15.33 E85Q 18% conversion of 10-desoxymethymycin, ratio of products neomethymycin to methymycin is 0.26. 8.1% conversion of substrate narbomycin Streptomyces venezuelae
1.14.15.33 E94Q no conversion of 10-desoxymethymycin. 5.4% conversion of substrate narbomycin Streptomyces venezuelae

Organism

EC Number Organism UniProt Comment Textmining
1.14.15.33 Streptomyces venezuelae O87605
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Substrates and Products (Substrate)

EC Number Substrates Comment Substrates Organism Products Comment (Products) Rev. Reac.
1.14.15.33 10-deoxymethymycin + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 i.e. YC-17. 40% conversion, ratio of products neomethymycin to methymycin is 0.81 Streptomyces venezuelae methymycin + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.33 10-deoxymethymycin + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 40% conversion, ratio of products neomethymycin to methymycin is 0.81 Streptomyces venezuelae neomethymycin + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O
-
 ?
1.14.15.33 narbomycin + 2 reduced ferredoxin [iron-sulfur] cluster + 2 H+ + O2 binding kinetics are biphasic and accurately described by a double exponential function revealing a fast first phase followed by a slow second one. The kinetic rate of the first binding step, accounting for about 50% of the reaction amplitude, is fast and protein-dependent. The rate of the second step is slow, about 13 per s Streptomyces venezuelae pikromycin + 2 oxidized ferredoxin [iron-sulfur] cluster + H2O 35% conversion ?