SYNTHESIS OF COBALAMIN-BIOTIN CONJUGATES THAT VARY IN THE POSITION OFCOBALAMIN COUPLING - EVALUATION OF COBALAMIN DERIVATIVE BINDING TO TRANSCOBALAMIN-II
Pm. Pathare et al., SYNTHESIS OF COBALAMIN-BIOTIN CONJUGATES THAT VARY IN THE POSITION OFCOBALAMIN COUPLING - EVALUATION OF COBALAMIN DERIVATIVE BINDING TO TRANSCOBALAMIN-II, Bioconjugate chemistry, 7(2), 1996, pp. 217-232
Six cobalamin-biotin conjugates have been prepared. The cobalamin-biot
in conjugates were prepared to evaluate the effect that the location o
f attachment had on the binding with transcobalamin II (TCII), the cob
alamin binding protein in plasma, and to evaluate their potential use
for in vitro and in vivo applications. This study focused only on the
effect of binding with TCII. To decrease the possibility of steric pro
blems in binding of the cobalamin conjugates with TCII, and biotin's b
inding with streptavidin or avidin, moieties of 11-18 atoms in length
were used as linkers. Four biotin conjugates were prepared which were
attached to the corrin ring of the cobalamin molecule (on b-, c-, d-,
and e-side chains). One conjugate was attached to the 5'-OH of the rib
ose moiety, and another conjugate was attached at the cobalt metal (in
place of the cyanide moiety of cyanocobalamin). Competitive binding s
tudies were conducted where various amounts of the cobalamin-biotin co
njugates and their precursor cobalamin derivatives competed with [Co-5
7]cyanocobalamin for binding of recombinant human TCII (rhTCII). Evalu
ation of cobalamin derivatives which were conjugated at the 5'-OH of r
ibose or the cobalt metal center indicated that conjugation at either
of these positions had Little effect on binding with rhTCII. However,
conjugates where the attachment was made on the corrin ring substituen
ts had a large variation in binding with rhTCII. Conjugates on the e-p
ropionamide side chain had little effect (relative affinity was equal
to or decreased less than a factor of 3) on binding with rhTCII, conju
gates of the b-isomer had decreased binding (relative affinity decreas
ed less than a factor of 10), conjugates of the d-propionamide had fur
ther decreased binding (relative affinity decreased between 44 and 69
times), and conjugates on the c-acetamide group had poor binding to rh
TCII (relative affinity decreased between 295 and 1160 times). The sig
nificance of the side chains on the corrin ring in providing specifici
ty and high-affinity binding with rhTCII is discussed.