Intermediate channeling between ATP sulfurylase and adenosine 5'-phosphosulfate kinase from rat chondrosarcoma.
Biosynthesis of the activated sulfate donor PAPS (3'-phosphoadenosine 5'-phosphosulfate) involves the sequential action of two enzyme activities. ATP sulfurylase catalyzes the formation of APS (adenosine 5'-phosphosulfate) from ATP and free sulfate, and APS is then phosphorylated by APS kinase to produce PAPS. Using rat chondrosarcoma ATP sulfurylase and APS kinase, a newly developed assay system, which permits measuring the accumulation of both APS and PAPS in the presence of both enzyme activities, produces a PAPS/APS ratio corresponding to a "channeling efficiency" of 96%. The velocity of the APS kinase reaction measured in the overall system with endogenously synthesized APS is 8-fold greater than that of the isolated kinase reaction using exogenous APS. Most conclusively, isotope dilution and enrichment experiments show that the APS intermediate does not equilibrate with APS in the bulk medium but remains largely bound in the rat enzyme system. In contrast, control experiments with a nonchanneled system containing a mixture of the sulfurylase and kinase isolated from Penicillium chrysogenum give the results expected for a nonchanneled pathway. These data indicate that APS is channeled between the active sites of ATP sulfurylase and APS kinase during the production of PAPS in rat chondrosarcoma.
Lyle
S
S
Department of Pediatrics, University of Chicago, Illinois 60637.
Ozeran
J D
JD
Stanczak
J
J
Westley
J
J
Schwartz
N B
NB
D000250:Adenosine Phosphosulfate / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D000818:Animals; D002813:Chondrosarcoma / Q000201:enzymology; D007700:Kinetics; D008956:Models, Chemical; D010724:Phosphoadenosine Phosphosulfate / Q000096:biosynthesis*; D010761:Phosphorus Radioisotopes; D017853:Phosphotransferases (Alcohol Group Acceptor) / Q000378:metabolism*; D051381:Rats; D013430:Sulfate Adenylyltransferase / Q000378:metabolism*; D013462:Sulfur Radioisotopes
D010761:Phosphorus Radioisotopes; D013462:Sulfur Radioisotopes; D010724:Phosphoadenosine Phosphosulfate; D000250:Adenosine Phosphosulfate; D000255:Adenosine Triphosphate; D017853:Phosphotransferases (Alcohol Group Acceptor); C019617:adenylylsulfate kinase; D013430:Sulfate Adenylyltransferase