Crystallization and X-ray analysis of native and selenomethionyl beta-mannanase Man5A from blue mussel, Mytilus edulis, expressed in Pichia pastoris.
The glycohydrolase family 5 beta-mannanase Man5A from Mytilus edulis has been expressed in Pichia pastoris and purified in a form suitable for X-ray crystallographic analysis. Crystals were grown by the hanging-drop technique at 293 K using polyethylene glycol 5000 monomethylether as precipitant and dioxane as additive. The crystals belong to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 61.8, b = 64.8, c = 90.7 A. Diffraction to 1.4 A resolution has been obtained at 100 K. Expression was also performed in the presence of selenomethionine. The incorporation of SeMet was estimated at 40% by amino-acid analysis and its presence in crystals was confirmed from the X-ray absorption scanning spectrum.
Xu
Bingze
B
Center for Surface Biotechnology, Uppsala University Biomedical Center, PO Box 577, SE-751 23, Uppsala, Sweden.
Muñoz I
Inés G
IG
Janson
Jan-Christer
JC
Ståhlberg
Jerry
J
D000818:Animals; D049872:Bivalvia / Q000737:chemistry / Q000201:enzymology*; D003001:Cloning, Molecular; D003460:Crystallization; D018360:Crystallography, X-Ray; D008361:Mannosidases / Q000096:biosynthesis / Q000737:chemistry* / Q000235:genetics; D010843:Pichia / Q000235:genetics; D011487:Protein Conformation; D011994:Recombinant Proteins / Q000737:chemistry; D012645:Selenomethionine / Q000378:metabolism; D044902:beta-Mannosidase
D011994:Recombinant Proteins; D012645:Selenomethionine; D008361:Mannosidases; D044902:beta-Mannosidase