Conformations of synthetic beta peptides in solid state and in aqueous solution: relation to toxicity in PC12 cells.

The secondary structures of peptides beta 25-35 (the active toxic fragment) and beta 35-25 (reverse sequence and non-toxic fragment), as well as of the amidated beta (25-35)-NH2 peptide were investigated in aqueous solution and in the solid state by means of Fourier-transformed infrared spectroscopy and circular dichroism spectroscopy. The conformations of the beta 25-35 and beta 35-25 in solid state were identical and contained mostly beta-sheet structures. In solid state the amidated beta (25-35)-NH2 peptide also contained mostly beta-sheet structures.

Thermal and pH stabilities of alkaline phosphatase from bovine intestinal mucosa: a FTIR study.

The inactivation of alkaline phosphatase (AP) from bovine intestinal mucosa caused by lowering the p2H from 10.4 to 5.4 or by increasing the temperature from 25 degrees C to 70 degrees C were not followed by significant FTIR changes, indicating that the native conformation of AP was preserved under these conditions.

Human pancreatic lipase. Importance of the hinge region between the two domains, as revealed by monoclonal antibodies.

Several monoclonal antibodies (mAbs) were prepared against human pancreatic lipase (HPL). Two enzyme-linked immunosorbent assay (ELISA) procedures were set up for screening hybridomas producing specific antibodies. Four mAbs (81-23, 146-40, 315-25, and 320-24) of the IgG1 isotype were found to react with HPL in both simple sandwich and double sandwich ELISAs, while mAb 248-31, of the IgG2b isotype, reacted only with HPL in a double sandwich ELISA.

Immunochemical studies of pancreatic colipase-lipase interaction employing immobilized synthetic peptides.

In view to study the possible participation of the sequence portions of colipase including or close to the free carboxyl groups at positions 15 and/or 72 to the binding with pancreatic lipase, we have used three synthetic peptides matching portions 8-16, 59-67 and 67-72 of the amino acid sequence. Polyclonal rabbit anticolipase immune serum, which cross-reacts with peptides in ELISA, was fractionated on columns of peptide coupled to Sepharose. Of the three fractions of antibodies, only that interacting with peptide 8-16 had the capacity to inhibit colipase-dependent lipase activity by specifically preventing the association of lipase with its protein cofactor previously bound to lipid.

Separation and characterization of the precursor and activated forms of porcine and human pancreatic colipase by reversed-phase liquid chromatography.

Reversed-phase liquid chromatography was used as an alternative method for the characterization of the precursor and activated forms of porcine and human pancreatic colipase. Using a Beckman Ultrasphere column with an increasing acetonitrile gradient in 0.1% trifluoroacetic acid, it was possible to obtain well-resolved separation of the precursor form of colipase (procolipase) from its trypsin-activated derivative.

Antigen specificity and cross-species reactivity of a monoclonal antibody (mAb 72.11) against porcine pancreatic procolipase.

We have studied the antigen specificity and cross-reactivity of a monoclonal antibody (mAb 72.11) of subclass IgG1, raised against the precursor form of porcine colipase (procolipase), whose epitope lies near the amino terminal region of the polypeptide. mAb 72.

Immunochemical determination of porcine pancreatic colipase: differentiation between procolipase and its trypsin-activated form.

A noncompetitive enzyme-linked immunosorbent assay (ELISA) has been developed for the quantitative determination of porcine pancreatic colipase. Calibration curves were established by coating polystyrene immunoplates with pure procolipase or its trypsin-activated derivative. Bound antigen was detected with antiporcine procolipase polyclonal antibodies.

Inhibitory properties and antigenic specificity of monoclonal antibodies to pancreatic colipase.

To understand the mechanism by which colipase acts as a protein cofactor for anchoring pancreatic lipase at triacylglycerol/water interface, we have used an immunochemical approach. Ten monoclonal antibodies (Mabs) against porcine pancreatic procolipase were produced. Purified immunoglobulins and Fab fragments were studied for their capacity to inhibit colipase-dependent lipase activity.

Production and characterization of four monoclonal antibodies against porcine pancreatic colipase.

Four monoclonal antibodies directed against porcine colipase have been generated by hybridization of myeloma cells with spleen cells of BALB/c immunized mice. Antibodies were screened by binding to immobilized colipase in a solid-phase assay. Monoclonal antibodies were purified by affinity chromatography on colipase coupled to Sepharose.