Premium
X‐ray diffraction analysis and in vitro characterization of the UAM2 protein from Oryza sativa
Author(s) -
Welner Ditte Hededam,
Tsai Alex Yi-Lin,
DeGiovanni Andy M.,
Scheller Henrik Vibe,
Adams Paul D.
Publication year - 2017
Publication title -
acta crystallographica section f
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.572
H-Index - 37
ISSN - 2053-230X
DOI - 10.1107/s2053230x17004587
Subject(s) - proteolysis , x ray crystallography , chemistry , size exclusion chromatography , diffraction , crystallography , protein crystallization , monomer , proteolytic enzymes , enzyme , biochemistry , crystallization , organic chemistry , physics , optics , polymer
The role of seemingly non‐enzymatic proteins in complexes interconverting UDP‐arabinopyranose and UDP‐arabinofuranose (UDP‐arabinosemutases; UAMs) in the plant cytosol remains unknown. To shed light on their function, crystallographic and functional studies of the seemingly non‐enzymatic UAM2 protein from Oryza sativa (OsUAM2) were undertaken. Here, X‐ray diffraction data are reported, as well as analysis of the oligomeric state in the crystal and in solution. OsUAM2 crystallizes readily but forms highly radiation‐sensitive crystals with limited diffraction power, requiring careful low‐dose vector data acquisition. Using size‐exclusion chromatography, it is shown that the protein is monomeric in solution. Finally, limited proteolysis was employed to demonstrate DTT‐enhanced proteolytic digestion, indicating the existence of at least one intramolecular disulfide bridge or, alternatively, a requirement for a structural metal ion.