A. Filipek et al., INTERACTION OF CALCYCLIN AND ITS CYANOGEN-BROMIDE FRAGMENTS WITH ANNEXIN-II AND GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE, International journal of biochemistry & cell biology, 27(11), 1995, pp. 1123-1131
The structural properties of calcyclin protein are quite well characte
rized but its function remains obscure. To help elucidate the biologic
al role of calcyclin we have performed the in vitro studies of the Ca2
+-dependent interaction of Ehrlich ascites tumor cells calcyclin and i
ts cyanogen bromide fragments with two potential calcyclin targets: an
nexin II and glyceraldehyde 3-phosphate dehydrogenase (GAPDH), The bin
ding of annexin II, evidenced by the reaction with I-125-calcyclin, wa
s found to be very weak and occurred only for intact calcyclin. On the
other hand the interaction between calcyclin and GAPDH was of high af
finity and could be assigned to the N-terminal region of calcyclin. In
tact calcyclin and its N-terminal fragment bound to GAPDH in the gel o
verlay and affinity chromatography assay. When examined in the presenc
e of a crosslinking agent the interaction resulted in the formation of
46K covalent adduct between calcyclin monomer and GAPDH subunit. Fluo
rescence of 5-iodoacetamido-fluorescein-labelled calcyclin was efficie
ntly quenched by GAPDH in the presence of Ca2+, Titration experiments
revealed the stoichiometry of one calcyclin monomer binding to each of
GAPDH subunits with a binding constant of 10(8) M(-1). The results of
this work suggest that the binding between calcyclin and GAPDH may ha
ve bearing on calcyclin function.