title: The hetero-hexameric nature of a chloroplast AAA+ FtsH protease contributes to its thermodynamic stability.
abstract: FtsH is an evolutionary conserved membrane-bound metalloprotease complex. While in most prokaryotes FtsH is encoded by a single gene, multiple FtsH genes are found in eukaryotes. Genetic and biochemical data suggest that the Arabidopsis chloroplast FtsH is a hetero-hexamer. This raises the question why photosynthetic organisms require a heteromeric complex, whereas in most bacteria a homomeric one is sufficient. To gain structural information of the possible complexes, the Arabidopsis FtsH2 (type B) and FtsH5 (type A) were modeled. An in silico study with mixed models of FtsH2/5 suggests that heteromeric hexamer structure with ratio of 4:2 is more likely to exists. Specifically, calculation of the buried surface area at the interfaces between neighboring subunits revealed that a hetero-complex should be thermodynamically more stable than a homo-hexamer, due to the presence of additional hydrophobic and hydrophilic interactions. To biochemically assess this model, we generated Arabidopsis transgenic plants, expressing epitope-tagged FtsH2 and immuno-purified the protein. Mass-spectrometry analysis showed that FtsH2 is associated with FtsH1, FtsH5 and FtsH8. Interestingly, we found that 'type B' subunits (FtsH2 and FtsH8) were 2-3 fold more abundant than 'type A' (FtsH1 and FtsH5). The biochemical data corroborate the in silico model and suggest that the thylakoid FtsH hexamer is composed of two 'type A' and four 'type B' subunits.
authors.affiliation: The Robert H. Smith Institute of Plant Sciences and Genetics in Agriculture, Hebrew University of Jerusalem, Rehovot, Israel.
authors.forename: Ofer
authors.initials: O
authors.lastname: Moldavski
authors.forename: Olga
authors.initials: O
authors.lastname: Levin-Kravets
authors.forename: Tamar
authors.initials: T
authors.lastname: Ziv
authors.forename: Zach
authors.initials: Z
authors.lastname: Adam
authors.forename: Gali
authors.initials: G
authors.lastname: Prag
mesh_terms: D049069:ATP-Dependent Proteases / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D000595:Amino Acid Sequence; D017360:Arabidopsis / Q000201:enzymology* / Q000235:genetics; D029681:Arabidopsis Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D002736:Chloroplasts / Q000201:enzymology*; D004795:Enzyme Stability; D000939:Epitopes / Q000378:metabolism; D013058:Mass Spectrometry; D008565:Membrane Proteins / Q000737:chemistry* / Q000302:isolation & purification / Q000378:metabolism; D008666:Metalloendopeptidases / Q000737:chemistry* / Q000378:metabolism; D045726:Metalloproteases / Q000737:chemistry* / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D009097:Multienzyme Complexes / Q000378:metabolism; D030821:Plants, Genetically Modified; D055503:Protein Multimerization; D021122:Protein Subunits / Q000737:chemistry / Q000302:isolation & purification / Q000378:metabolism; D021381:Protein Transport; D011993:Recombinant Fusion Proteins / Q000302:isolation & purification / Q000378:metabolism; D013816:Thermodynamics; D020524:Thylakoids / Q000201:enzymology
chemical_list: D049069:ATP-Dependent Proteases; D029681:Arabidopsis Proteins; D000939:Epitopes; D008565:Membrane Proteins; D008666:Metalloendopeptidases; D045726:Metalloproteases; D009097:Multienzyme Complexes; D021122:Protein Subunits; D011993:Recombinant Fusion Proteins; C547215:FtsH5 protein, Arabidopsis; C402651:AT2G30950 protein, Arabidopsis; C114583:m-AAA proteases