PubMed:10085236 JSONTXT

{"project":"PMID_GLOBAL","denotations":[{"id":"T1","span":{"begin":919,"end":922},"obj":"DiseaseOrPhenotypicFeature"},{"id":"T3","span":{"begin":1412,"end":1421},"obj":"DiseaseOrPhenotypicFeature"}],"attributes":[{"id":"A1","pred":"mondo_id","subj":"T1","obj":"0004974"},{"id":"A2","pred":"mondo_id","subj":"T1","obj":"0022673"},{"id":"A3","pred":"mondo_id","subj":"T3","obj":"0000605"}],"text":"ATPase activity associated with the magnesium-protoporphyrin IX chelatase enzyme of Synechocystis PCC6803: evidence for ATP hydrolysis during Mg2+ insertion, and the MgATP-dependent interaction of the ChlI and ChlD subunits.\nInsertion of Mg2+ into protoporphyrin IX catalysed by the three-subunit enzyme magnesium-protoporphyrin IX chelatase (Mg chelatase) is thought to be a two-step reaction, consisting of activation followed by Mg2+ chelation. The activation step requires ATP and two of the subunits, ChlI and ChlD (I and D respectively), and it has been speculated that this step results in the formation of an I-D-ATP complex. The subsequent step, in which Mg2+ is inserted into protoporphyrin, also requires ATP and the third subunit, H, in addition to ATP-activated I-D complex. In the present study, we examine the interaction of the I and D subunits of the Mg chelatase from the cyanobacterium Synechocystis PCC 6803. We demonstrate the purification of an I-D complex, and show that ATP and Mg2+ are absolute requirements for the formation of this complex, probably as MgATP. However, ATP may be replaced by the slowly hydrolysable analogue, adenosine 5'-[gamma-thio]triphosphate, and, to a minor extent, by ADP and the non-hydrolysable ATP analogue, adenosine 5'-[beta,gamma-imido]triphosphate, all of which suggests that ATP hydrolysis is not necessary for the formation of the ChlI-ChlD complex. A sensitive continuous assay was used to detect ATPase activity during Mg2+ chelation, and it was found that the maximum rate of ATP hydrolysis coincided with the maximum rate of Mg2+ insertion. The rate of ATP hydrolysis depended on factors that determined the rate of Mg2+ chelation, such as increasing the concentration of the H subunit and the concentration of protoporphyrin. Thus ATP hydrolysis has been identified as an absolute requirement for the chelation step. The I subunit possessed strong ATPase activity when assayed on its own, whereas the D subunit had no detectable activity, and when the I and D subunits were assayed in combination, the ATPase activity of the I subunit was repressed."}

{"project":"CyanoBase","denotations":[{"id":"T1","span":{"begin":201,"end":205},"obj":"protein"},{"id":"T2","span":{"begin":210,"end":214},"obj":"protein"},{"id":"T3","span":{"begin":844,"end":845},"obj":"protein"},{"id":"T4","span":{"begin":850,"end":851},"obj":"protein"},{"id":"T5","span":{"begin":967,"end":968},"obj":"protein"},{"id":"T6","span":{"begin":969,"end":970},"obj":"protein"},{"id":"T7","span":{"begin":1391,"end":1395},"obj":"protein"},{"id":"T8","span":{"begin":1396,"end":1400},"obj":"protein"},{"id":"T9","span":{"begin":1740,"end":1741},"obj":"protein"},{"id":"T10","span":{"begin":1886,"end":1887},"obj":"protein"},{"id":"T11","span":{"begin":1966,"end":1967},"obj":"protein"},{"id":"T12","span":{"begin":2017,"end":2018},"obj":"protein"},{"id":"T13","span":{"begin":2023,"end":2024},"obj":"protein"},{"id":"T14","span":{"begin":2090,"end":2091},"obj":"protein"}],"text":"ATPase activity associated with the magnesium-protoporphyrin IX chelatase enzyme of Synechocystis PCC6803: evidence for ATP hydrolysis during Mg2+ insertion, and the MgATP-dependent interaction of the ChlI and ChlD subunits.\nInsertion of Mg2+ into protoporphyrin IX catalysed by the three-subunit enzyme magnesium-protoporphyrin IX chelatase (Mg chelatase) is thought to be a two-step reaction, consisting of activation followed by Mg2+ chelation. The activation step requires ATP and two of the subunits, ChlI and ChlD (I and D respectively), and it has been speculated that this step results in the formation of an I-D-ATP complex. The subsequent step, in which Mg2+ is inserted into protoporphyrin, also requires ATP and the third subunit, H, in addition to ATP-activated I-D complex. In the present study, we examine the interaction of the I and D subunits of the Mg chelatase from the cyanobacterium Synechocystis PCC 6803. We demonstrate the purification of an I-D complex, and show that ATP and Mg2+ are absolute requirements for the formation of this complex, probably as MgATP. However, ATP may be replaced by the slowly hydrolysable analogue, adenosine 5'-[gamma-thio]triphosphate, and, to a minor extent, by ADP and the non-hydrolysable ATP analogue, adenosine 5'-[beta,gamma-imido]triphosphate, all of which suggests that ATP hydrolysis is not necessary for the formation of the ChlI-ChlD complex. A sensitive continuous assay was used to detect ATPase activity during Mg2+ chelation, and it was found that the maximum rate of ATP hydrolysis coincided with the maximum rate of Mg2+ insertion. The rate of ATP hydrolysis depended on factors that determined the rate of Mg2+ chelation, such as increasing the concentration of the H subunit and the concentration of protoporphyrin. Thus ATP hydrolysis has been identified as an absolute requirement for the chelation step. The I subunit possessed strong ATPase activity when assayed on its own, whereas the D subunit had no detectable activity, and when the I and D subunits were assayed in combination, the ATPase activity of the I subunit was repressed."}