Open AccessMolecular Properties of Lipoprotein Lipase
Author(s) -
OLIVECRONA Thomas,
BENGTSSON Gunilla,
OSBORNE James C.
Publication year - 1982
Publication title -
european journal of biochemistry
Language(s) - English
Resource type - Journals
eISSN - 1432-1033
pISSN - 0014-2956
DOI - 10.1111/j.1432-1033.1982.tb06640.x
Subject(s) - guanidinium chloride , chemistry , sedimentation equilibrium , lipase , gel permeation chromatography , trypsin , chromatography , lipoprotein lipase , partial specific volume , monomer , ultracentrifuge , enzyme , biochemistry , organic chemistry , polymer
The monomer molecular size of bovine lipoprotein lipase was evaluated by sedimentation equilibrium measurements and by gel permeation chromatography in 6 M guanidinium chloride. To establish molecular weight unequivocally we determined the partial specific volume ( v̄ ) experimentally. This was done by analyzing equilibrium concentration profiles from analytical ultracentrifugation in 6 M guanidinium chloride using buffers made up in H 2 O and 2 H 2 O. The combined results gave a v̄ of 0.71 ± 0.007 ml/g and a molecular weight of 41700 ± 1000 for monomeric bovine lipoprotein lipase. This value did not change upon mild tryptic digestion; the elution volume upon gel permeation chromatography in 6 M guanidinium chloride was also unaffected by treatment with trypsin. Sedimentation equilibrium measurements of the trypsin‐treated material in the presence of reducing agents gave limiting molecular weights of 19000 and 23000, demonstrating that mild trypsin digestion cleaved lipoprotein lipase into two polypeptide chains of similar size held together by disulfide bonds. Mild trypsin digestion also resulted in a loss of secondary structure as determined by circular dichroic measurements. Discussion centers around the correlation between these effects of trypsin on the molecular properties of lipoprotein lipase and the previously reported effects on the kinetic properties of the enzyme.