PubMed Health⌕ Search

Biomedical subjects

C Chapus

Publications and source records attributed to C Chapus.

45 records · Page 3Linked to original sources
PubMed1981

Amino acid sequence of horse colipase B.

The complete sequence of the 96 residues composing horse colipase B has been determined by automated analysis of the intact protein, of two CNBr peptides and two tryptic peptides arising, respectively, from the citraconylated chain and from the unreduced protein. The single histidine of the protein is located at position 29 as in horse colipase A. His86, present in the C-terminal region of the pig cofactor and supposed to play a role in the folding molecule, is not conserved in horse B. Large pieces of the pig and horse B chains were found to be identical or very similar, especially the N-terminal sequence and the central segment Ala49-Cys65 including the three tyrosines of the molecule. The four lysines and the ten half cystines are also conserved.

Amino Acid Sequence↗
PubMed1981

Stabilization of the C-terminal part of pig and horse colipase by carboxypeptidase and trypsin inhibitors.

Pig and horse colipases have been purified by a common procedure using trypsin and carboxypeptidase inhibitors as stabilizers. Two forms of pig colipase were identified: a predominant A1 form with about 103-105 residues, and a minor slightly degraded A2 form in which the last two C-terminal residues, Asp and Ser, were lacking. This type of degradation is considerably slowed down by carboxypeptidase inhibitors. A total of four forms of the horse cofactor were characterized: two (A1 and B1) were probably isocolipases which differed by only a few substitutions. Both contained the same number of residues (about 96), an N-terminal valine and an Arg-Ser-Glu-(Glx)1,2-ArgC-terminal sequence. A2 and B2 were slightly degraded forms probably resulting from tryptic cleavage of the Arg-Ser bond in the above sequence. The presence of methionine in the horse cofactor allowed fragmentation by cyanogen bromide. The C-terminal fragment was composed of 16 or 17 residues and contained no histidine. The single histidine of horse B1 was found in the intermediary fragment between Met-18 and Met-(n-17). These data show that the C-terminal parts of both pig and horse colipases are still more exposed to proteolytic degradations than the N-terminal parts. Preliminary attempts to crystallize B1 were carried out.

Animals↗
PubMed1978

Adsorption and activation of pancreatic lipase at interfaces.

The first step of the lipase-catalyzed hydrolysis of insoluble long chain triglycerides is the adsorption of the enzyme to the interface. This adsorption, which is spontaneous when the interface is hydrophobic, is hindered by bile salts. Under these conditions, a small protein cofactor designated colipase adsorbs first and then anchors lipase at the interface. Interfacial adsorption enhances lipase activity, due, at least in part, to an acceleration of the rate-limiting deacylation step of the reaction. In this respect, lipase appears to be a most interesting model of an enzyme being activated by the presence of a lipid. The 3 steps of the heterogeneous catalysis induced by lipase, interfacial adsorption, interfacial activation and catalysis proper are under the control, respectively, of a serine hydroxyl group, a carboxyl and a histidine imidazole.

Adsorption↗
PubMed1976

Mechanism of pancreatic lipase action. 1. Interfacial activation of pancreatic lipase.

Hydrolysis of dissolved p-nitrophenyl acetate by pancreatic lipase follows the classical acyl enzyme pathway already proposed for other esterases. Kinetic parameters of the hydrolysis have been determined. The turnover rate of the reaction is many orders of magnitude slower than that for the natural emulsified substrates. Nevertheless, several arguments are in favor of the specificity of this hydrolysis: (1) triacetin, which resembles the usual substrates for the enzyme, is also hydrolyzed very slowly in solution; (2) dissolved triacetin and tripropionin are competitive inhibitors for the p-nitrophenyl acetate hydrolysis; (3) the same chemical structural features which are required in the case of emulsified substrates are also necessary to promote hydrolysis of dissolved p-nitrophenyl esters. This suggests that the same active site (or part of the same active site) is responsible for hydrolysis of both p-nitrophenyy acetate and specific emulsified substrates. Since deacylation is the rate-limiting step in the catalysis of p-nitrophenyl acetate, the intermediate acetyl enzyme can be isolated by trapping it at pH 5.0. Kinetic competence of this intermediate has been demonstrated. Hydrolysis by pancreatic lipase of dissolved monomeric p-nitrophenyl acetate and triacetin is considerably enhanced (100- to 500-fold) by various interfaces. This suggests that at least the deacylation step, which is rate limiting in absence of interface, is accelerated by the presence of inert interfaces. Siliconized glass beads were directly shown to accelerate the deacylation of isolated [3H]acetyl lipase by at least a hundred times. This step does not directly involve the ester substrate.Thus, it is suggested that a part of the activation of lipase at interfaces may be due to a conformational change resulting from adsorption.

Acetates↗
PubMed1976

Mechanism of pancreatic lipase action. 2. Catalytic properties of modified lipases.

Reaction of lipase with diethyl pyrocarbonate results in the modification of three histidine residues. One is highly reactive, although without affecting the activity, while the two others react more slowly with a concomitant loss of activity on both dissolved and emulsified substrates. As previously shown, lipase can also be modified either by reaction of five carboxyl groups with carbodiimide (5N-lipase) or by esterification of one serine residue with diethyl p-nitrophenyl phosphate (DP-lipase). In the three cases, the activity on emulsified substrates is abolished. The modification of histidine residues results also in a loss of activity on dissolved substrates, suggesting that the essential histidine is at (or close to) the active site. The ability of lipase to be adsorbed on siliconized glass beads is not impaired in this reaction. By contrast, 5N-lipase is still able to hydrolyze dissolved monomeric substrates and to adsorb on siliconized glass beands. Therefore, the essential carboxyl group is assumed to play an important role in the interfacial activation. Finally, since DP-lipase is still fully active on dissolved p-nitrophenyl acetate, the serine residue, which has been previously suggested to be the acylable one, is more likely implicated in the recognition and the binding to interfaces, as confirmed by the inability of DP-lipase to be adsorbed on siliconized glass beads.

Diethyl Pyrocarbonate↗