Cloning, expression, purification, crystallization and initial crystallographic analysis of the lysine-biosynthesis LysX protein from Thermus thermophilus HB8.
The gene encoding LysX, an essential component of the lysine-biosynthesis pathway in Thermus thermophilus (molecular weight approximately equal 31,000 Da), was cloned and expressed and the purified protein was crystallized by the hanging-drop vapour-diffusion technique in two different space groups, C2 (unit-cell parameters a = 124.7, b = 51.4, c = 103.6 A, beta = 122.8 degrees ) and R3 (a = b = 122.6, c = 97.6 A). Crystals improved by macroseeding diffracted to beyond 2.3 and 3 A resolution for the C2 and R3 crystal forms, respectively. Complete diffraction data sets were collected for the C2 and R3 crystal forms at 2.5 and 3.1 A resolution, respectively. Crystals of selenomethionine-containing LysX protein were obtained by cross-microseeding, using the native microcrystals as a seed. Structure determination is now in progress.
Vassylyeva
Marina N
MN
Genomic Sciences Center, RIKEN Yokohama Institute, 1-7-22 Suehiro-cho, Tsurumi, Yokohama 230-0045, Japan.
Sakai
Hiroaki
H
Matsuura
Takanori
T
Sekine
Shun-ichi
S
Nishiyama
Makoto
M
Terada
Takaho
T
Shirouzu
Mikako
M
Kuramitsu
Seiki
S
Vassylyev
Dmitry G
DG
Yokoyama
Shigeyuki
S
D001426:Bacterial Proteins / Q000737:chemistry*; D003001:Cloning, Molecular / Q000379:methods; D003460:Crystallization / Q000379:methods*; D018360:Crystallography, X-Ray; D008239:Lysine / Q000096:biosynthesis / Q000737:chemistry*; D016963:Thermus thermophilus / Q000201:enzymology*
D001426:Bacterial Proteins; C477978:LysX protein, T thermophilus; D008239:Lysine