PURIFICATION AND CHARACTERIZATION OF RECOMBINANT MOUSE GROWTH-HORMONEBINDING-PROTEIN PRODUCED IN THE BACULOVIRUS EXPRESSION SYSTEM

Sf21 insect cells were infected with recombinant baculovirus containin g cDNA for the entire coding region of the mouse growth hormone bindin g protein (mGHBP). Recombinant (r) mGHBP was expressed at a yield of 1 7.3 mg/liter/3 days. The molecular size (M(r)) of the rmGHBP was appro ximately 33,000 as estimated by SDS-PAGE. Amino-terminal sequence anal ysis of the recombinant protein yielded two sequences: one identical t o amino acids 1-15 and another corresponding to amino acids 14-21 of t he GHR/GHBP. Western blot analysis revealed that this is the same M(r) as that of one of the two major M(r) forms of serum mGHBP. Deglycosyl ation of serum mGHBP and recombinant mGHBP caused a shift in the molec ular size of both proteins to that expected after removal of all N-lin ked carbohydrates. Binding characteristics of the recombinant mGHBP to mouse growth hormone were similar to those for serum GHBP. Scatchard analysis showed an equilibrium association constant (K-a) for rmGHBP o f 3.8 x 10(8) +/- 0.6 x 10(8) M(-1) (mean +/- SEM, n = 3) and K-a of 9 .2 x 10(8) +/- 2.0 x 10(8) M(-1) (mean +/- SEM, n = 3) for the serum m GHBP. In conclusion, this expression system should allow a production of relatively large quantities of mGHBP suitable for physiological stu dies on the role of this protein. (C) 1996 Academic Press, Inc.