title: Catalytic turnover triggers exchange of subunits of the magnesium chelatase AAA+ motor unit.
abstract: The ATP-dependent insertion of Mg(2+) into protoporphyrin IX is the first committed step in the chlorophyll biosynthetic pathway. The reaction is catalyzed by magnesium chelatase, which consists of three gene products: BchI, BchD, and BchH. The BchI and BchD subunits belong to the family of AAA+ proteins (ATPases associated with various cellular activities) and form a two-ring complex with six BchI subunits in one layer and six BchD subunits in the other layer. This BchID complex is a two-layered trimer of dimers with the ATP binding site located at the interface between two neighboring BchI subunits. ATP hydrolysis by the BchID motor unit fuels the insertion of Mg(2+) into the porphyrin by the BchH subunit. In the present study, we explored mutations that were originally identified in semidominant barley (Hordeum vulgare L.) mutants. The resulting recombinant BchI proteins have marginal ATPase activity and cannot contribute to magnesium chelatase activity although they apparently form structurally correct complexes with BchD. Mixing experiments with modified and wild-type BchI in various combinations showed that an exchange of BchI subunits in magnesium chelatase occurs during the catalytic cycle, which indicates that dissociation of the complex may be part of the reaction mechanism related to product release. Mixing experiments also showed that more than three functional interfaces in the BchI ring structure are required for magnesium chelatase activity.
authors.affiliation: Carlsberg Laboratory, Gamle Carlsberg Vej 10, 1799 Copenhagen V, Denmark.
authors.forename: Joakim
authors.initials: J
authors.lastname: Lundqvist
authors.forename: Ilka
authors.initials: I
authors.lastname: Braumann
authors.forename: Marzena
authors.initials: M
authors.lastname: Kurowska
authors.forename: André H
authors.initials: AH
authors.lastname: Müller
authors.forename: Mats
authors.initials: M
authors.lastname: Hansson
mesh_terms: D000251:Adenosine Triphosphatases / Q000737:chemistry / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000378:metabolism; D055162:Biocatalysis; D002734:Chlorophyll / Q000378:metabolism; D001467:Hordeum / Q000201:enzymology*; D008190:Lyases / Q000737:chemistry / Q000378:metabolism* / Q000648:ultrastructure; D020409:Molecular Motor Proteins / Q000378:metabolism*; D009154:Mutation / Q000235:genetics; D055503:Protein Multimerization; D021122:Protein Subunits / Q000737:chemistry / Q000378:metabolism*; D013053:Spectrophotometry
chemical_list: D020409:Molecular Motor Proteins; D021122:Protein Subunits; D002734:Chlorophyll; D000255:Adenosine Triphosphate; D000251:Adenosine Triphosphatases; D008190:Lyases; C086788:magnesium chelatase
keywords: AAA; ATPases; Biosynthesis; Chelatase; Chlorophyll Biosynthesis; Magnesium; Magnesium Chelatase; Plant Biochemistry; Porphyrin; Protoporphyrin