We reconstituted ovine (o) LH alpha from its amino- and carboxyl-termi
nal fragments obtained as follows. oLH alpha was nicked at Arg(46)-Ser
(47) with Arg-C protease. Nicked oLH alpha disulfide bonds were broken
by sulfitolysis, and its N-terminal peptide and C-terminal glycopepti
de were separated by Sephacryl S-200 chromatography. Both fragments we
re mixed, reduced, and reoxidized. Reoxidation products were chromatog
raphed on Sephacryl S-200, and an alpha-monomer fraction was recovered
. The putative nicked alpha-monomer fraction was reassociated with nat
ive oLH beta, and the resulting oLH derivative was isolated by S-200 c
hromatography with a reduced yield of 11% (intact subunits yield, 67%
oLH). This preparation was 2.6% as active as oLH in a LH receptor bind
ing assay. Two additional oLH derivatives were prepared Cleavage at al
pha Arg(46)-Ser(47) alone, followed by reassociation with native oLH b
eta, produced Arg-C-nicked oLH alpha:oLH beta (14% yield) that was 3.3
% as active as native oLH. Reduction-reoxidation of Arg-C-nicked oLH a
lpha followed by reassociation with oLH beta produced reduced reoxidiz
ed-Arg-C-nicked oLH alpha:oLH beta (11% yield) that was 1.8% as active
as oLH. These results indicated that the nicked oLH alpha monomer had
been reconstituted from its N- and C-terminal fragments.