Use of novel selenomethionine-resistant yeast to produce selenomethionyl protein suitable for structural analysis.
Yeast is widely used to determine the tertiary structure of eukaryotic proteins, because of its ability to undergo post-translational modifications such as glycosylation. A mutant lacking S-adenosylmethionine synthesis has been reported as a suitable host for producing selenomethionine derivatives, which can help solve phase problems in protein crystallography. However, the mutant required external addition of S-adenosylmethionine for cell proliferation. Here, a selenomethionine-resistant Pichia pastoris mutant that showed S-adenosylmethionine autotrophy was isolated. Human lysozyme expressed by the mutant under the control of constitutive promoter contained selenomethionine at 65% occupancy, sufficient for use as a selenomethionine derivative for single-wavelength anomalous dispersion phasing.
Kitajima
Toshihiko
T
Research Center for Medical Glycoscience, National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, Japan.
Yagi
Emi
E
Kubota
Tomomi
T
Chiba
Yasunori
Y
Nishikawa
Satoshi
S
Jigami
Yoshifumi
Y
D009113:Muramidase / Q000096:biosynthesis / Q000737:chemistry / Q000235:genetics; D010843:Pichia / Q000302:isolation & purification / Q000378:metabolism*; D011994:Recombinant Proteins / Q000096:biosynthesis* / Q000737:chemistry / Q000235:genetics; D012436:S-Adenosylmethionine / Q000378:metabolism; D012645:Selenomethionine / Q000378:metabolism*
D011994:Recombinant Proteins; D012436:S-Adenosylmethionine; D012645:Selenomethionine; D009113:Muramidase