Determination of Kinetics and the Crystal Structure of a Novel Type 2 Isopentenyl Diphosphate: Dimethylallyl Diphosphate Isomerase from Streptococcus pneumoniae.

Chembiochem. 2014 Jun 6. doi: 10.1002/cbic.201402046. [Epub ahead of print]

Determination of Kinetics and the Crystal Structure of a Novel Type 2 Isopentenyl Diphosphate: Dimethylallyl Diphosphate Isomerase from Streptococcus pneumoniae.

de Ruyck J1, Janczak MW, Neti SS, Rothman SC, Schubert HL, Cornish RM, Matagne A, Wouters J, Poulter CD.

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Abstract

Isopentenyl diphosphate isomerase (IDI) is a key enzyme in the isoprenoid biosynthetic pathway and is required for all organisms that synthesize isoprenoid metabolites from mevalonate. Type 1 IDI (IDI-1) is a metalloprotein that is found in eukaryotes, whereas the type 2 isoform (IDI-2) is a flavoenzyme found in bacteria that is completely absent from human. IDI-2 from the pathogenic bacterium Streptococcus pneumoniae was recombinantly expressed in Escherichia coli. Steady-state kinetic studies of the enzyme indicated that FMNH2 (KM =0.3 μM) bound before isopentenyl diphosphate (KM =40 μM) in an ordered binding mechanism. An X-ray crystal structure at 1.4 Å resolution was obtained for the holoenzyme in the closed conformation with a reduced flavin cofactor and two sulfate ions in the active site. These results helped to further approach the enzymatic mechanism of IDI-2 and, thus, open new possibilities for the rational design of antibacterial compounds against sequence-similar and structure-related pathogens such as Enterococcus faecalis or Staphylococcus aureus.

© 2014 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

KEYWORDS:

X-ray structures; bisubstrates; flavoproteins; isoprenoids; steady-state kinetics

PMID: 24910111 [PubMed - as supplied by publisher]