Kinetic mechanism of adenosine 5'-phosphosulphate kinase from rat chondrosarcoma.

Abstract

Biosynthesis of the activated sulphate donor adenosine 3'-phosphate 5'-phosphosulphate (PAPS) involves the sequential action of two enzyme activities. ATP-sulphurylase catalyses the formation of APS (adenosine 5'-phosphosulphate) from ATP and free sulphate, and APS is then phosphorylated by APS kinase to produce PAPS. Initial-velocity patterns for rat chondrosarcoma APS kinase indicate a single-displacement formal mechanism with KmAPS 76 nM and KmATP = 24 microM. Inhibition studies using analogues of substrates and products were carried out to determine the reaction mechanism. An analogue of PAPS, adenosine 3'-phosphate 5'-[beta-methylene]phosphosulphate, exhibited competitive inhibition with APS and non-competitive inhibition with ATP. An analogue of APS, adenosine 5'-[beta-methylene]phosphosulphate was also competitive with APS and non-competitive with ATP. Adenosine 5'-[beta gamma-imido]triphosphate showed competitive inhibition with respect to ATP and produced mixed-type inhibition, with a pronounced intercept effect and a small slope effect, with respect to APS. These results are in accord with the formulation of the predominant pathway as a steady-state ordered mechanism with APS as the leading substrate and PAPS as the final product released.

• Authors: S Lyle, D H Geller, K Ng, J Stanczak, J Westley, N B Schwartz
• Journal: The Biochemical journal (Biochem J) Vol. 301 ( Pt 2) Pg. 355-9 (Jul 15 1994) ISSN: 0264-6021 [Print] England
• PMID: 8042977 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't, Research Support, U.S. Gov't, P.H.S.)

Chemical References

• Adenosine Phosphosulfate
• Adenosine Triphosphate
• Phosphotransferases (Alcohol Group Acceptor)
• adenylylsulfate kinase
• Sulfate Adenylyltransferase
• Phosphoric Monoester Hydrolases
• 3'-phosphoadenosine-5'-phosphosulfate phosphohydrolase

Topics

• Adenosine Phosphosulfate (metabolism)
• Adenosine Triphosphate (metabolism)
• Animals
• Binding, Competitive
• Chondrosarcoma (enzymology)
• Kinetics
• Phosphoric Monoester Hydrolases (metabolism)
• Phosphorylation
• Phosphotransferases (Alcohol Group Acceptor) (antagonists & inhibitors, metabolism)
• Rats
• Sulfate Adenylyltransferase (metabolism)