Analyses of carbohydrate binding property of lectin-chaperone calreticulin.
Calreticulin (CRT) is a soluble molecular chaperone of the endoplasmic reticulum. It is a lectin that promotes the folding of proteins carrying N-linked glycans. Recent investigations have revealed that glucosylated high-mannose-type glycans are employed as key elements in this process. Here, we performed quantitative analyses of the interaction of CRT with various disaccharides, including fluorine-substituted analogues using a quartz-crystal microbalance (QCM). These experiments revealed the weak affinity of 2- and 3-fluoroglucose derivatives. On the other hand, 6-fluoroglucose derivatives exhibited a significant affinity, indicating that the role of 6-position of OH is less significant for binding to CRT. We also characterized binding epitope of the Glcalpha1-3Man(alpha)Me to CRT by saturation transfer difference (STD) NMR spectroscopy. It is proposed that 2-, 3-, and 4-positions of Glc and 3-, 4-, and 6-positions of Man are in close contact with CRT binding pocket, while 6-position of Glc and 2-position of Man are not. These finding are in excellent agreement with our QCM experiment.
Tatami
Atsushi
A
RIKEN (The Institute of Physical and Chemical Research), 2-1 Hirosawa, Wako, Saitama 351-0198, Japan.
Hon
Yung-Son
YS
Matsuo
Ichiro
I
Takatani
Maki
M
Koshino
Hiroyuki
H
Ito
Yukishige
Y
D000818:Animals; D037282:Calreticulin / Q000737:chemistry* / Q000235:genetics / Q000378:metabolism; D004186:Disaccharidases / Q000737:chemistry; D000939:Epitopes; D005461:Fluorine; D005978:Glutathione / Q000235:genetics; D006801:Humans; D037102:Lectins / Q000378:metabolism*; D009682:Magnetic Resonance Spectroscopy; D018832:Molecular Chaperones / Q000737:chemistry* / Q000378:metabolism; D011485:Protein Binding; D011993:Recombinant Fusion Proteins / Q000737:chemistry / Q000235:genetics
D037282:Calreticulin; D000939:Epitopes; D037102:Lectins; D018832:Molecular Chaperones; D011993:Recombinant Fusion Proteins; D005461:Fluorine; D004186:Disaccharidases; D005978:Glutathione