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Structural studies on KijD1, a sugar C ‐3′‐methyltransferase
Author(s) -
Dow Garrett T.,
Thoden James B.,
Holden Hazel M.
Publication year - 2016
Publication title -
protein science
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 3.353
H-Index - 175
eISSN - 1469-896X
pISSN - 0961-8368
DOI - 10.1002/pro.3034
Subject(s) - stereochemistry , protein quaternary structure , chemistry , enzyme , dimer , methyltransferase , biochemistry , o methyltransferase , mutagenesis , transferase , cofactor , mutation , organic chemistry , dna , gene , methylation , protein subunit
Abstract Kijanimicin is an antitumor antibiotic isolated from Actinomadura kijaniata . It is composed of three distinct moieties: a pentacyclic core, a monosaccharide referred to as d ‐kijanose, and a tetrasaccharide chain composed of l ‐digitoxose units. d ‐Kijanose is a highly unusual nitro‐containing tetradeoxysugar, which requires at least ten enzymes for its production. Here we describe a structural analysis of one of these enzymes, namely KijD1, which functions as a C ‐3′‐methyltransferase using S ‐adenosylmethionine as its cofactor. For this investigation, two ternary complexes of KijD1, determined in the presence of S ‐adenosylhomocysteine (SAH) and dTDP or SAH and dTDP‐3‐amino‐2,3,6‐trideoxy‐4‐keto‐3‐methyl‐ d ‐glucose, were solved to 1.7 or 1.6 Å resolution, respectively. Unexpectedly, these structures, as well as additional biochemical analyses, demonstrated that the quaternary structure of KijD1 is a dimer. Indeed, this is in sharp contrast to that previously observed for the sugar C ‐3′‐methyltransferase isolated from Micromonospora chalcea . By the judicious use of site‐directed mutagenesis, it was possible to convert the dimeric form of KijD1 into a monomeric version. The quaternary structure of KijD1 could not have been deduced based solely on bioinformatics approaches, and thus this investigation highlights the continuing need for experimental validation.