Preliminary X-ray crystallographic analysis of the secreted chorismate mutase from Mycobacterium tuberculosis: a tricky crystallization problem solved

Journal Article · Mon May 01 04:00:00 EDT 2006 · Acta Crystallographica. Section F

DOI:https://doi.org/10.1107/S1744309106012036· OSTI ID:22356324

Krengel, Ute ^[1]; Dey, Raja ^[1]; Sasso, Severin ^[2]; Ökvist, Mats ^[1]; Ramakrishnan, Chandra; Kast, Peter ^[2]

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1. Department of Chemistry and Bioscience, Chalmers University of Technology, PO Box 462, SE-40530 Göteborg (Sweden)
2. Laboratory of Organic Chemistry, ETH Zurich, CH-8093 Zurich (Switzerland)

A method is presented that allowed the diffraction limit of crystals of the secreted chorismate mutase from M. tuberculosis to be improved from approximately 3.5 to 1.3 Å. To obtain large well diffracting crystals, it was critical to initiate crystallization at higher precipitant concentration and then transfer the drops to lower precipitant concentrations within 5–15 min. Chorismate mutase catalyzes the conversion of chorismate to prephenate in the biosynthesis of the aromatic amino acids tyrosine and phenylalanine in bacteria, fungi and plants. Here, the crystallization of the unusual secreted chorismate mutase from Mycobacterium tuberculosis (encoded by Rv1885c), a 37.2 kDa dimeric protein belonging to the AroQ{sub γ} subclass of mutases, is reported. Crystal optimization was non-trivial and is discussed in detail. To obtain crystals of sufficient quality, it was critical to initiate crystallization at higher precipitant concentration and then transfer the drops to lower precipitant concentrations within 5–15 min, in an adaptation of a previously described technique [Saridakis & Chayen (2000 ▶), Protein Sci.9, 755–757]. As a result of the optimization, diffraction improved from 3.5 to 1.3 Å resolution. The crystals belong to space group P2{sub 1}, with unit-cell parameters a = 42.6, b = 72.6, c = 62.0 Å, β = 104.5°. The asymmetric unit contains one biological dimer, with 167 amino acids per protomer. A soak with a transition-state analogue is also described.

• OSTI ID: 22356324
• Journal Information: Acta Crystallographica. Section F, Journal Name: Acta Crystallographica. Section F Journal Issue: Pt 5 Vol. 62; ISSN ACSFCL; ISSN 1744-3091
• Country of Publication: United Kingdom
• Language: English

Related Subjects

75 CONDENSED MATTER PHYSICS
SUPERCONDUCTIVITY AND SUPERFLUIDITY
CONVERSION
CRYSTALLIZATION
CRYSTALS
DIFFRACTION
DIMERS
NUCLEATION
OPTIMIZATION
PROTEINS
RESOLUTION
SPACE GROUPS