The activation of porcine pancreatic lipase by cis-unsaturated fatty acids.
In the presence of taurodeoxycholate, cis-unsaturated fatty acids increase porcine pancreatic lipase activity 15-fold at pH 7.5. This effect is saturable with a low proportion of fatty acid to substrate. The overall angle of the fatty acid, the position of its double bond and the presence of a carboxyl group were critical factors in whether the fatty acid effectively increased lipase activity. When the substrate is emulsified by taurodeoxycholate, the pH optimum for lipase ranges from 6.2 to 7.0. In the presence of cis-unsaturated fatty acids, the overall activity of lipase increases, the pH optimum shifts, and the pH-activity curve becomes biphasic, with one optimum around pH 7.7, and the other around pH 8.8. Fluorescence studies indicate that fatty acids bind near aromatic residues in lipase, particularly tryptophan. Using the fluorescent fatty acid cis-parinaric acid, it was determined that multiple binding sites are present with Kd values of approx. 10(-6) M. Far-UV circular dichroism (CD) studies indicate that in addition to a high affinity fatty acid binding site with a Kd of approx. 10(-6) M, there is also a low affinity binding site with a Kd of approx. 10(-4) M. The far-UV CD data also show that cis-unsaturated fatty acids change the conformation of lipase. It is calculated that the percentage of alpha helix decreases, and the amount of beta sheet and beta turn structure increases. Because the three-dimensional crystal structure of lipase is known, a model is proposed to describe how cis-unsaturated fatty acids increase lipase activity.
van Kuiken
B A
BA
Department of Molecular Genetics, Biochemistry and Microbiology, University of Cincinnati College of Medicine, OH 45267.
Behnke
W D
WD
D000818:Animals; D001665:Binding Sites; D002942:Circular Dichroism; D003089:Colipases / Q000378:metabolism; D004305:Dose-Response Relationship, Drug; D004789:Enzyme Activation / Q000187:drug effects; D005231:Fatty Acids, Unsaturated / Q000494:pharmacology*; D005453:Fluorescence; D006863:Hydrogen-Ion Concentration; D019787:Linoleic Acid; D008041:Linoleic Acids / Q000494:pharmacology; D008049:Lipase / Q000378:metabolism*; D019301:Oleic Acid; D009829:Oleic Acids / Q000494:pharmacology; D010179:Pancreas / Q000201:enzymology*; D011487:Protein Conformation; D013552:Swine
D003089:Colipases; D005231:Fatty Acids, Unsaturated; D008041:Linoleic Acids; D009829:Oleic Acids; D019301:Oleic Acid; D019787:Linoleic Acid; D008049:Lipase