ATPase activity of magnesium chelatase subunit I is required to maintain subunit D in vivo.
During biosynthesis of chlorophyll, Mg(2+) is inserted into protoporphyrin IX by magnesium chelatase. This enzyme consists of three different subunits of approximately 40, 70 and 140 kDa. Seven barley mutants deficient in the 40 kDa magnesium chelatase subunit were analysed and it was found that this subunit is essential for the maintenance of the 70 kDa subunit, but not the 140 kDa subunit. The 40 kDa subunit has been shown to belong to the family of proteins called "ATPases associated with various cellular activities", known to form ring-shaped oligomeric complexes working as molecular chaperones. Three of the seven barley mutants are semidominant mis-sense mutations leading to changes of conserved amino acid residues in the 40 kDa protein. Using the Rhodobacter capsulatus 40 and 70 kDa magnesium chelatase subunits we have analysed the effect of these mutations. Although having no ATPase activity, the deficient 40 kDa subunit could still associate with the 70 kDa protein. The binding was dependent on Mg(2+) and ATP or ADP. Our study demonstrates that the 40 kDa subunit functions as a chaperon that is essential for the survival of the 70 kDa subunit in vivo. We conclude that the ATPase activity of the 40 kDa subunit is essential for this function and that binding between the two subunits is not sufficient to maintain the 70 kDa subunit in the cell. The ATPase deficient 40 kDa proteins fail to participate in chelation in a step after the association of the 40 and 70 kDa subunits. This step presumably involves a conformational change of the complex in response to ATP hydrolysis.
Lake
Vanessa
V
Department of Biological Science, Macquarie University, North Ryde, Australia.
Olsson
Ulf
U
Willows
Robert D
RD
Hansson
Mats
M
D000251:Adenosine Triphosphatases / Q000378:metabolism*; D001467:Hordeum / Q000201:enzymology / Q000235:genetics; D008190:Lyases / Q000032:analysis / Q000235:genetics / Q000378:metabolism*; D009154:Mutation; D021122:Protein Subunits / Q000032:analysis / Q000235:genetics / Q000378:metabolism; D012333:RNA, Messenger / Q000032:analysis
D021122:Protein Subunits; D012333:RNA, Messenger; D000251:Adenosine Triphosphatases; D008190:Lyases; C086788:magnesium chelatase