1.55A-resolution structure of ent-copalyl diphosphate synthase and exploration of general acid function by site-directed mutagenesis.

ABSTRACT:

Background

The diterpene cyclase ent-copalyl diphosphate synthase (CPS) catalyzes the first committed step in the biosynthesis of gibberellins. The previously reported 2.25Å resolution crystal structure of CPS complexed with (S)-15-aza-14,15-dihydrogeranylgeranyl thiolodiphosphate (1) established the αβγ domain architecture, but ambiguities regarding substrate analog binding remained.

Method

Use of crystallization additives yielded CPS crystals diffracting to 1.55Å resolution. Additionally, active site residues that hydrogen bond with D379, either directly or through hydrogen bonded water molecules, were probed by mutagenesis.

Results

This work clarifies structure-function relationships that were ambiguous in the lower resolution structure. Well-defined positions for

SUBMITTER: Koksal M 

PROVIDER: S-EPMC3859867 | biostudies-literature | 2014 Jan

REPOSITORIES: biostudies-literature