Molecular basis for semidominance of missense mutations in the XANTHA-H (42-kDa) subunit of magnesium chelatase.
During biosynthesis of bacteriochlorophyll or chlorophyll, three protein subunits of 140, 70, and 42 kDa interact to insert Mg2+ into protoporphyrin IX. The semidominant Chlorina-125, -157, and -161 mutants in barley are deficient in this step and accumulate protoporphyrin IX after feeding on 5-aminolevulinate. Chlorina-125, -157, and -161 are allelic to the recessive xantha-h mutants and contain G559A, G806A, and C271T mutations, respectively. These mutations cause single amino acid substitutions in residues that are conserved in all known primary structures of the 42-kDa subunit. In vitro complementation and reconstitution of Mg-chelatase activity show that the 42-kDa subunits are defective in the semidominant Chlorina mutants. A mutated protein is maintained in the Chlorina plastids, unlike in the xantha-h plastids. Heterozygous Chlorina seedlings have 25-50% of the Mg-chelatase activity of wild-type seedlings. Codominant expression of active and inactive 42-kDa subunits in heterozygous Chlorina seedlings is likely to produce two types of heterodimers between the strongly interacting 42-kDa and 70-kDa subunits. Reduced Mg-chelatase activity is explained by the capacity of heterodimers consisting of mutated 42-kDa and wild-type 70-kDa protein to bind to the 140-kDa subunit. The 42-kDa subunit is similar to chaperones that refold denatured polypeptides with respect to its ATP-to-ADP exchange activity and its ability to generate ATPase activity with the 70-kDa subunit. We hypothesize that the association of the 42-kDa subunit with the 70-kDa subunit allows them to form a specific complex with the 140-kDa subunit and that this complex inserts Mg2+ into protoporphyrin IX.
Hansson
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A
Department of Biochemistry, Center for Chemistry and Chemical Engineering, Lund University, P.O. Box 124, S-221 00 Lund, Sweden.
Kannangara
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CG
von Wettstein
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Hansson
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D000595:Amino Acid Sequence; D019943:Amino Acid Substitution; D001419:Bacteria / Q000201:enzymology / Q000235:genetics; D002736:Chloroplasts / Q000201:enzymology; D005816:Genetic Complementation Test; D006579:Heterozygote; D001467:Hordeum / Q000201:enzymology* / Q000235:genetics*; D007700:Kinetics; D008190:Lyases / Q000737:chemistry / Q000235:genetics* / Q000378:metabolism; D046911:Macromolecular Substances; D008969:Molecular Sequence Data; D020125:Mutation, Missense; D018515:Plant Leaves; D010944:Plants / Q000201:enzymology / Q000235:genetics; D017354:Point Mutation; D016415:Sequence Alignment; D017386:Sequence Homology, Amino Acid