Cloning, expression, purification and crystallization of dihydrodipicolinate synthase from Agrobacterium tumefaciens
Sarah C Atkinson1, Con Dogovski, Renwick C J Dobson
1Department of Biochemistry, La Trobe Institute for Molecular Science, La Trobe University, Melbourne, Victoria 3086, Australia.
Abstract:
Dihydrodipicolinate synthase (DHDPS) catalyzes the first committed step of the lysine-biosynthesis pathway in bacteria, plants and some fungi. This study describes the cloning, expression, purification and crystallization of DHDPS (NP_354047.1) from the plant pathogen Agrobacterium tumefaciens (AgT-DHDPS). Enzyme-kinetics studies demonstrate that AgT-DHDPS possesses DHDPS activity in vitro. Crystals of AgT-DHDPS were grown in the unliganded form and in forms with substrate bound and with substrate plus allosteric inhibitor (lysine) bound. X-ray diffraction data sets were subsequently collected to a maximum resolution of 1.40 Å. Determination of the structure with and without substrate and inhibitor will offer insight into the design of novel pesticide agents.
