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Title: A single point mutation changes the crystallization behavior of Mycoplasma arthritidis-derived mitogen

Abstract

The mutagenesis, crystallization and preliminary crystallographic analysis of M. arthritidis-derived mitogen is described. Mycoplasma arthritidis-derived mitogen (MAM) functions as a conventional superantigen (SAg). Although recombinant MAM has been crystallized by the hanging-drop vapour-diffusion method, the crystals diffracted poorly to only 5.0 Å resolution, with large unit-cell parameters a = 163.8, b = 93.0, c = 210.9 Å, β = 93.7° in the monoclinic space group P2{sub 1}. Unit-cell content analysis revealed that as many as 24 molecules could be present in the asymmetric unit. Systematic alanine mutagenesis was applied in order to search for mutants that give crystals of better quality. Two mutants, L50A and K201A, were crystallized under the same conditions as wild-type MAM (MAM{sub wt}). Crystals of the L50A mutant are isomorphous with those of MAM{sub wt}, while a new crystal form was obtained for the K201 mutant, belonging to the cubic space group P4{sub 1}32 with unit-cell parameters a = b = c = 181.9 Å. Diffraction data were collected to 3.6 and 2.8 Å resolution from crystals of the MAM L50A and K201A mutants, respectively. Molecular-replacement calculations suggest the presence of two molecules in the asymmetric unit for the MAM K201A mutant crystal, resulting in amore » V{sub M} of 5.0 Å Da{sup −1} and a solvent content of 75%. An interpretable electron-density map for the MAM K201A mutant crystal was produced using the molecular-replacement method.« less

Authors:
; ;  [1];  [1];  [2]
  1. Wadsworth Center, New York State Department of Health, Empire State Plaza, PO Box 509, Albany, New York 12201-0509 (United States)
  2. (United States)
Publication Date:
OSTI Identifier:
22356292
Resource Type:
Journal Article
Resource Relation:
Journal Name: Acta Crystallographica. Section F; Journal Volume: 62; Journal Issue: Pt 3; Other Information: PMCID: PMC2197180; PMID: 16511311; PUBLISHER-ID: fw5068; OAI: oai:pubmedcentral.nih.gov:2197180; Copyright (c) International Union of Crystallography 2006; Country of input: International Atomic Energy Agency (IAEA)
Country of Publication:
United Kingdom
Language:
English
Subject:
75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND SUPERFLUIDITY; CHEMICAL ANALYSIS; CRYSTALLIZATION; CRYSTALS; DENSITY; DIFFRACTION; DIFFUSION; ELECTRON DENSITY; MOLECULES; PLASMA; RESOLUTION; SOLVENTS; SPACE GROUPS

Citation Formats

Guo, Yi, Li, Zhong, Van Vranken, Sandra J., Li, Hongmin, E-mail: lih@wadsworth.org, and Department of Biomedical Sciences, School of Public Health, University at Albany, State University of New York, Empire State Plaza, PO Box 509, Albany, New York 12201-0509. A single point mutation changes the crystallization behavior of Mycoplasma arthritidis-derived mitogen. United Kingdom: N. p., 2006. Web. doi:10.1107/S1744309106003691.
Guo, Yi, Li, Zhong, Van Vranken, Sandra J., Li, Hongmin, E-mail: lih@wadsworth.org, & Department of Biomedical Sciences, School of Public Health, University at Albany, State University of New York, Empire State Plaza, PO Box 509, Albany, New York 12201-0509. A single point mutation changes the crystallization behavior of Mycoplasma arthritidis-derived mitogen. United Kingdom. doi:10.1107/S1744309106003691.
Guo, Yi, Li, Zhong, Van Vranken, Sandra J., Li, Hongmin, E-mail: lih@wadsworth.org, and Department of Biomedical Sciences, School of Public Health, University at Albany, State University of New York, Empire State Plaza, PO Box 509, Albany, New York 12201-0509. Wed . "A single point mutation changes the crystallization behavior of Mycoplasma arthritidis-derived mitogen". United Kingdom. doi:10.1107/S1744309106003691.
@article{osti_22356292,
title = {A single point mutation changes the crystallization behavior of Mycoplasma arthritidis-derived mitogen},
author = {Guo, Yi and Li, Zhong and Van Vranken, Sandra J. and Li, Hongmin, E-mail: lih@wadsworth.org and Department of Biomedical Sciences, School of Public Health, University at Albany, State University of New York, Empire State Plaza, PO Box 509, Albany, New York 12201-0509},
abstractNote = {The mutagenesis, crystallization and preliminary crystallographic analysis of M. arthritidis-derived mitogen is described. Mycoplasma arthritidis-derived mitogen (MAM) functions as a conventional superantigen (SAg). Although recombinant MAM has been crystallized by the hanging-drop vapour-diffusion method, the crystals diffracted poorly to only 5.0 Å resolution, with large unit-cell parameters a = 163.8, b = 93.0, c = 210.9 Å, β = 93.7° in the monoclinic space group P2{sub 1}. Unit-cell content analysis revealed that as many as 24 molecules could be present in the asymmetric unit. Systematic alanine mutagenesis was applied in order to search for mutants that give crystals of better quality. Two mutants, L50A and K201A, were crystallized under the same conditions as wild-type MAM (MAM{sub wt}). Crystals of the L50A mutant are isomorphous with those of MAM{sub wt}, while a new crystal form was obtained for the K201 mutant, belonging to the cubic space group P4{sub 1}32 with unit-cell parameters a = b = c = 181.9 Å. Diffraction data were collected to 3.6 and 2.8 Å resolution from crystals of the MAM L50A and K201A mutants, respectively. Molecular-replacement calculations suggest the presence of two molecules in the asymmetric unit for the MAM K201A mutant crystal, resulting in a V{sub M} of 5.0 Å Da{sup −1} and a solvent content of 75%. An interpretable electron-density map for the MAM K201A mutant crystal was produced using the molecular-replacement method.},
doi = {10.1107/S1744309106003691},
journal = {Acta Crystallographica. Section F},
number = Pt 3,
volume = 62,
place = {United Kingdom},
year = {Wed Mar 01 00:00:00 EST 2006},
month = {Wed Mar 01 00:00:00 EST 2006}
}