The human phosphoadenosine-phosphosulfate synthase (PAPS) system is a bifunctional enzyme (fusion product of two catalytic activities). In a first step, sulfate adenylyltransferase catalyses the formation of adenosine 5′-phosphosulfate (APS) from ATP and inorganic sulfate. The second step is catalysed by the adenylylsulfate kinase portion of 3′-phosphoadenosine 5′-phosphosulfate (PAPS) synthase, which involves the formation of PAPS from enzyme-bound APS and ATP.
MPLPKRSIEPVHVARSVYQQDELQSVELETVTNTTLTNIIRQLSSLSKHAEDVFGELARD
VGNIGDRANSLQARIDRLAIKVTQLDSTVEEVPLTDITRKKAFKSAKVFDQQIFSRATMP
APMMDTYAQCDKPPPLDKLNVYRDDGKDGLKFYTDPNYFFELWRQEMLKDTERVMHDKGK
KLNRPRQDGGAGGAAGRGNKKQKTKIRVPHNTREQQRQRALVHGETLMPNNVIYRTPNSM
VNEEAGYGNTVKPYNMLSHLENNSNVTKQTAVAIADMGVYDTRPPRPNSIELNRSYQSEQ
IDGSTYEQLPPQMGNQYAATFGNGMGGPQQMHMQHQQMYDAGMYQSHALYGQTGQGVMSP
EPIYGPGTPSRNKPRPSQPPPAPPSNGSGGGTPTASNANTPTRGRSMSTSRDALPPPPPV
PDVISPMSGMNGVNSGHMAAKLLGRANSSSGAGSPNSVQNANDMVMTQLSNTFHSIGMTG
NQLNSLSDLPPPPPVPDQHSPKMSPPNAAPPPPPPPPPVEEGMGSGNQHTLRPHQILPKS
LANGEMQQPGQQNGVPHIVAPKKMLPPFHDPRNDLMKAIRDGITLRKVEKSEQKEIERNA
APLDVASILARRVAIELSESEDSDSEDDSEGWMEPNETSA650
| 9668121 | Molecular cloning, expression, and characterization of human bifunctional 3'-phosphoadenosine 5'-phosphosulfate synthase and its functional domains.Venkatachalam KV, Akita H, Strott CA | The universal sulfonate donor, 3'-phosphoadenosine 5'-phosphosulfate (PAPS), is synthesized by the concerted action of ATP sulfurylase and adenosine 5'-phosphosulfate (APS) kinase, which in animals are fused into a bifunctional protein. The cDNA for human PAPS synthase (hPAPSS) along with polymerase chain reaction products corresponding to several NH2- and COOH-terminal fragments were cloned and expressed in COS-1 cells. A 1-268-amino acid fragment expressed APS kinase activity, whereas a 220-623 fragment evinced ATP sulfurylase activity. The 1-268 fragment and full-length hPAPSS (1-623) exhibited hyperbolic responses against APS substrate with equivalent Km values (0.6 and 0.4 microM, respectively). The 1-268 fragment demonstrated Michaelis-Menten kinetics against ATP as substrate (Km 0.26 mM); however, full-length hPAPSS exhibited a sigmoidal response (apparent Km 1.5 mM) suggesting cooperative binding. Catalytic efficiency (Vmax/Km) of the 1-268 fragment was 64-fold higher than full-length hPAPSS for ATP. The kinetic data suggest that the COOH-terminal domain of hPAPSS exerts a regulatory role over APS kinase activity located in the NH2-terminal domain of this bifunctional protein. In addition, the 1-268 fragment and full-length hPAPSS were overexpressed in Escherichia coli and column purified. Purified full-length hPAPSS, in contrast to the COS-1 cell-expressed cDNA construct, exhibited a hyperbolic response curve against ATP suggesting that hPAPSS is perhaps modified in vivo. | 1998 |
Venkatachalam K V , Akita H , Strott C A . Molecular Cloning, Expression, and Characterization of Human Bifunctional 3"-Phosphoadenosine 5"-Phosphosulfate Synthase and Its Functional Domains[J]. Journal of Biological Chemistry, 1998, 273(30):19311-19320.