Identification and crystallization of a protease-resistant core of calnexin that retains biological activity.
Calnexin is a molecular chaperone that facilitates folding of glycoproteins in the endoplasmic reticulum (ER). The cloned lumenal domain of canine calnexin, cnxDeltaTMC, retains its biological activity without the transmembrane and cytosolic region. For the purpose of structure determination we generated a crystallizable core by mild proteolysis and identified its termini by N-terminal sequencing and molecular mass determination. A truncated gene was cloned accordingly. Its product, cnxDeltaN25C15, was purified to apparent homogeneity and crystallized. This truncated variant remains biologically active as shown by its binding to monoglucosylated oligosaccharides and functional interaction with ERp57. A heavy atom derivative was identified.
Hahn
M
M
Biotechnology Research Institute, 6100 Royalmount Avenue, Montreal, H4P 2R2, Canada.
Borisova
S
S
Schrag
J D
JD
Tessier
D C
DC
Zapun
A
A
Tom
R
R
Kamen
A A
AA
Bergeron
J J
JJ
Thomas
D Y
DY
Cygler
M
M
D000595:Amino Acid Sequence; D000818:Animals; D002135:Calcium-Binding Proteins / Q000737:chemistry*; D037281:Calnexin; D003001:Cloning, Molecular; D003460:Crystallization; D004285:Dogs; D019286:Endopeptidase K / Q000378:metabolism; D010450:Endopeptidases / Q000378:metabolism*; D006023:Glycoproteins / Q000378:metabolism; D006360:Heat-Shock Proteins / Q000378:metabolism; D007535:Isomerases / Q000378:metabolism; D018832:Molecular Chaperones / Q000737:chemistry; D008969:Molecular Sequence Data; D009844:Oligosaccharides / Q000378:metabolism; D010446:Peptide Fragments / Q000737:chemistry*; D011485:Protein Binding; D011994:Recombinant Proteins / Q000737:chemistry; D017421:Sequence Analysis; D014357:Trypsin / Q000378:metabolism
D002135:Calcium-Binding Proteins; D006023:Glycoproteins; D006360:Heat-Shock Proteins; D018832:Molecular Chaperones; D009844:Oligosaccharides; D010446:Peptide Fragments; D011994:Recombinant Proteins; D037281:Calnexin; D010450:Endopeptidases; D014357:Trypsin; D019286:Endopeptidase K; D007535:Isomerases