Improvement of the quality of lumazine synthase crystals by protein engineering.
Icosahedral macromolecules have a wide spectrum of potential nanotechnological applications, the success of which relies on the level of accuracy at which the molecular structure is known. Lumazine synthase from Bacillus subtilis forms a 150 A icosahedral capsid consisting of 60 subunits and crystallizes in space group P6(3)22 or C2. However, the quality of these crystals is poor and structural information is only available at 2.4 A resolution. As classical strategies for growing better diffracting crystals have so far failed, protein engineering has been employed in order to improve the overexpression and purification of the molecule as well as to obtain new crystal forms. Two cysteines were replaced to bypass misfolding problems and a charged surface residue was replaced to force different molecular packings. The mutant protein crystallizes in space group R3, with unit-cell parameters a = b = 313.02, c = 365.77 A, alpha = beta = 90.0, gamma = 120 degrees , and diffracts to 1.6 A resolution.
Rodríguez-Fernández
Lidia
L
Department of Chemistry, Technische Universität München, Lichtenbergstrasse 4, 85747 Garching, Germany.
López-Jaramillo
F Javier
FJ
Bacher
Adelbert
A
Fischer
Markus
M
Weinkauf
Sevil
S
D001412:Bacillus subtilis / Q000201:enzymology / Q000235:genetics; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000592:standards; D003460:Crystallization / Q000379:methods / Q000592:standards; D009097:Multienzyme Complexes / Q000737:chemistry / Q000235:genetics / Q000592:standards*; D016297:Mutagenesis, Site-Directed / Q000379:methods / Q000592:standards; D015202:Protein Engineering / Q000379:methods / Q000592:standards*; D012258:Riboflavin Synthase / Q000737:chemistry / Q000235:genetics / Q000592:standards
D001426:Bacterial Proteins; D009097:Multienzyme Complexes; C044822:6,7-dimethyl-8-ribityllumazine synthase; D012258:Riboflavin Synthase