Investigation of two glycosylated forms of bile-salt-dependent lipase in human pancreatic juice.
Pure human pancreatic bile-salt-dependent lipase, devoid of its oncofetal glycoform [Mas, E., Abouakil, N., Roudani, S., Miralles, F., Guy-Crotte., O., Figarella, C., Escribano, M. J. & Lombardo, D. (1993) Biochem. J. 289, 609-615], was analyzed on immobilized concanavalin A (ConA). Two variants were separated: an unabsorbed ConA-unreactive fraction; and an absorbed ConA-reactive fraction. Carbohydrate compositions of ConA-reactive and ConA-unreactive fractions were not significantly different, and analysis of 3H-labelled oligosaccharides liberated from these fractions on the ConA-Sepharose column indicated that the fractionation of the bile-salt-dependent lipase on this column depends upon oligosaccharide structures. The activity of the ConA-reactive fraction was however much lower, independent of the substrate (4-nitrophenyl hexanoate or cholesteryl esters), than that of the ConA-unreactive fraction. Therefore, catalytic constants for the hydrolysis of 4-nitrophenyl hexanoate were determined; both fractions had quite similar Km, while the kcat for the ConA-unreactive fraction was 3-4-fold higher than that of the ConA-reactive fraction. ConA-reactive and ConA-unreactive fractions were shown to have slightly different molecular masses and different amino acid compositions. Cleavage patterns after cyanogen bromide treatment of the ConA-reactive and ConA-unreactive fractions suggested that the ConA-reactive (high Mr form) and ConA-unreactive (low Mr form) forms could be different isoforms of the bile-salt-dependent lipase secreted by the human pancreas.
Mas
E
E
INSERM U 260, Faculté de Médecine Marseille, France.
Franc
J L
JL
Lecestre
D
D
Crotte
C
C
Lombardo
D
D
Sadoulet
M O
MO
D000596:Amino Acids / Q000032:analysis; D000818:Animals; D002241:Carbohydrates / Q000032:analysis; D002352:Carrier Proteins / Q000378:metabolism; D003208:Concanavalin A / Q000737:chemistry; D006023:Glycoproteins / Q000378:metabolism*; D006801:Humans; D007700:Kinetics; D008049:Lipase / Q000737:chemistry / Q000378:metabolism*; D051379:Mice; D010189:Pancreatic Juice / Q000201:enzymology*; D002787:Sterol Esterase
D000596:Amino Acids; D002241:Carbohydrates; D002352:Carrier Proteins; D006023:Glycoproteins; D003208:Concanavalin A; C054784:bile salt-stimulated lipase; D002787:Sterol Esterase; C054621:CEL protein, human; D008049:Lipase