Structure analysis of geranyl pyrophosphate methyltransferase and the proposed reaction mechanism of SAM-dependentC-methylation

Abstract

In the typical isoprenoid-biosynthesis pathway, condensation of the universal C(5)-unit precursors isopentenyl pyrophosphate (IPP) and dimethylallyl pyrophosphate (DMAPP) occurs via the common intermediates prenyl pyrophosphates (C(10)-C(20)). The diversity of isoprenoids reflects differences in chain length, cyclization and further additional modification after cyclization. In contrast, the biosynthesis of 2-methylisonorneol (2-MIB), which is responsible for taste and odour problems in drinking water, is unique in that it primes the enzymatic methylation of geranyl pyrophosphate (GPP) before cyclization, which is catalyzed by an S-adenosyl-L-methionine-dependent methyltransferase (GPPMT). The substrate of GPPMT contains a nonconjugated olefin and the reaction mechanism is expected to be similar to that of the steroid methyltransferase (SMT) family. Here, structural analysis of GPPMT in complex with its cofactor and substrate revealed the mechanisms of substrate recognition and possible enzymatic reaction. Using the structures of these complexes, methyl-group transfer and the subsequent proton-abstraction mechanism are discussed. GPPMT and SMTs contain a conserved glutamate residue that is likely to play a role as a general base. Comparison with the reaction mechanism of the mycolic acid cyclopropane synthase (MACS) family also supports this result. This enzyme represented here is the first model of the enzymatic C-methylation of a nonconjugated olefin in the isoprenoid-biosynthesis pathway. In addition, an elaborate system to avoid methylation of incorrect substrates is proposed.

Structural and Molecular Basis of the Catalytic Mechanism of Geranyl Pyrophosphate C6‐Methyltransferase: Creation of an Unprecedented Farnesyl Pyrophosphate C6‐Methyltransferase

“…Despite the low sequence similarity,the overall structure of BezA is similar to other structurereported type Imethyltransferases,such as GPPC2MT (3VC2 and 4F86, amino acid sequence identity,19.8 and 21.2 %; root mean square deviation [RMSD],1.51 and 1.15 for and 115 Ca atoms,r espectively) and rebeccamycin sugar 4'-Omethyltransferase RebM (3BUS,i dentity 24.5 %; RMSD 1.13 for 132 Ca atoms;F igure S6). [4,[11][12][13][14][15] Theoverall structures of apo-BezA and BezA-SAH were nearly identical. TheR MSD value for 243 Ca atoms was 0.353 .H owever,t he length of the aZ1 helix differed between them (Figure 2a,b).…”

Structural Insights into the Catalytic Mechanism of Synechocystis Magnesium Protoporphyrin IX O-Methyltransferase (ChlM)

“…6A). Among them five catalyze N-methylation (42,46,48,51,52), two catalyze C-methylation (49,50), two catalyze the conversion of SAM to carboxy-SAM (43,44,53), one is a hydroxylase (47), and the only O-methyltransferase DhpI methylates the phosphonic acid group (45). Interestingly, like SyChlM, DhpI has an unusual insertion that encompasses a helix called the "capping helix" (topologically equivalent to ␣G in SyChlM) (Fig.…”

Structural and Molecular Basis of the Catalytic Mechanism of Geranyl Pyrophosphate C6‐Methyltransferase: Creation of an Unprecedented Farnesyl Pyrophosphate C6‐Methyltransferase

“…Thes tructures and reaction mechanisms of GPPC2MT from two different species, Streptomyces coelicolor (ScGPPC2MT,3 VC1, 3VC2, also known as SCO7701) and Streptomyces lasalocidi (SlGPPC2MT,4 F84, 4F85, 4F86), have been previously studied. [4,11] Determination of the crystal structures of these GPPC2MT enzymes binding with as ubstrate (or its analogue) and ac ofactor, and their subsequent biochemical analyses,led to aproposed molecular basis of the GPP methylation reaction (Figure 1c). First, the electrophilic attack of the reactive methyl group of SAM on the C2,C3 double bond of GPP forms aC 3c arbocation intermediate.…”