ArylamineN-Acetyltransferase Responsible for Acetylation of 2-Aminophenols inStreptomyces griseus

Abstract

An arylamine N-acetyltransferase (NAT) responsible for the N acetylation of exogenous 3-amino-4-hydroxybenzoic acid in Streptomyces griseus was identified and characterized. This enzyme was distinct from other eukaryotic and bacterial NATs in that it acetylated various 2-aminophenol derivatives more effectively than it acetylated 5-aminosalicylic acid, and thus it may be involved in the metabolism of xenobiotic compounds.

GriC and GriD Constitute a Carboxylic Acid Reductase Involved in Grixazone Biosynthesis in Streptomyces griseus

“…GriG is nonessential for grixazone biosynthesis but apparently contributes the uptake of 3,4-AHBA excreted and/or leaked from the cells (this study). 3,4-AcAHBA is produced from 3,4-AHBA by a cytosolic enzyme, SgNAT, encoded by natA in S. griseus [9]. SgNAT is also responsible for N-acetylation of 3,4-AHBAL to produce 3,4-AcAHBAL, but other acetyltransferase(s) must be involved in the N-acetylation (this study).…”

“…A natA null mutant (DnatA) of S. griseus was constructed previously [9]. S. griseus strains were grown at 30°C in YPD medium (0.2% yeast extract, 0.4% Bacto peptone, 0.5% NaCl, 0.2% MgSO 4 · 7H 2 O, 1.0% glucose, and 0.5% glycine, pH 7.2) or at 26.5°C in standard minimal medium [SMM; 0.9% glucose, 0.9% asparagine, 0.2% (NH 4 ) 2 SO 4 , 0.24% Trizma base, 0.1% NaCl, 0.05% K 2 SO 4 , 0.02% MgSO 4 · 7H 2 O, 0.01% CaCl 2 , 2.5 mM KH 2 PO 4 , and 1.0% trace element solution [10], pH 7.2].…”

“…SMM-0.25 contained a low concentration (0.25 mM) of KH 2 PO 4 to allow expression of the grixazone biosynthesis genes, because the gri genes are expressed under phosphate-depleted conditions [4]. Because 3,4-AcAHBA is a shunt product derived from 3,4-AHBA by an arylamine N-acetyltransferase (SgNAT) encoded by natA in S. griseus [9], these observations implied that both mutants accumulated 3,4-AHBA. Note that both 3,4-AHBA and 3,4-AcAHBA were never detected in the culture broth of the wild-type S. griseus cells under grixazone-producing conditions [6].…”

“…4, mutant griG::Km r [pIJ702-griG] converted 3,4-AHBA to 3,4-AcAHBA much more efficiently than mutant griG::Km r [pIJ702-melC1Ј]. Because 3,4-AcAHBA is produced from 3,4-AHBA by the cytosolic SgNAT [9] and it was very unlikely that GriG increases the enzyme activity of SgNAT, these results suggested that mutant griG::Km r [pIJ702-griG] strain was more efficient in the uptake of 3,4-AHBA (and/or the excretion of 3,4-AcAHBA) than mutant griG::Km r [pIJ702-melC1Ј].…”

“…Mutant D natA, which is deficient in the ability of Nacetylation of 3,4-AHBA [9], was adopted as the host for the bioconversion. In the bioconversion in which an aryl carboxylic acid was added at a final concentration of 10 mM, the most efficient substrate was 3,4-AHBA.…”