Showing papers by "Benedikt Westermann published in 1996"

The nucleotide exchange factor MGE exerts a key function in the ATP-dependent cycle of mt-Hsp70-Tim44 interaction driving mitochondrial protein import.

Abstract: Import of preproteins into the mitochondrial matrix is driven by the ATP-dependent interaction of mt-Hsp70 with the peripheral inner membrane import protein Tim44 and the preprotein in transit. We show that Mge1p, a co-chaperone of mt-Hsp70, plays a key role in the ATP-dependent import reaction cycle in yeast. Our data suggest a cycle in which the mt-Hsp70-Tim44 complex forms with ATP: Mge1p promotes assembly of the complex in the presence of ATP. Hydrolysis of ATP by mt-Hsp70 occurs in complex with Tim44. Mge1p is then required for the dissociation of the ADP form of mt-Hsp70 from Tim44 after release of inorganic phosphate but before release of ADP. ATP hydrolysis and complex dissociation are accompanied by tight binding of mt-Hsp70 to the preprotein in transit. Subsequently, the release of mt-Hsp70 from the polypeptide chain is triggered by Mge1p which promotes release of ADP from mt-Hsp70. Rebinding of ATP to mt-Hsp70 completes the reaction cycle.

Role of the mitochondrial DnaJ homolog Mdj1p as a chaperone for mitochondrially synthesized and imported proteins.

Abstract: Mdj1p,aDnaJhomologinthemitochondriaofSaccharomycescerevisiae,isinvolvedinthefoldingofproteins in the mitochondrial matrix. In this capacity, Mdj1p cooperates with mitochondrial Hsp70 (mt-Hsp70). Here, we analyzed the role of Mdj1p as a chaperone for newly synthesized proteins encoded by mitochondrial DNA and for nucleus-encoded proteins as they enter the mitochondrial matrix. A series of conditional mutants of mdj1wasconstructed.MutationsinthevariousfunctionaldomainsledtoapartiallossofMdj1pfunction.The mutant Mdj1 proteins were defective in protecting the tester protein firefly luciferase against heat-induced aggregationinisolatedmitochondria.Themitochondriallyencodedvar1proteinshowedenhancedaggregation after synthesis in mdj1 mutant mitochondria. Mdj1p and mt-Hsp70 were found in a complex with nascent polypeptide chains on mitochondrial ribosomes. Mdj1p was not found to interact with translocation intermediates of imported proteins spanning the two membranes and exposing short segments into the matrix, in accordancewiththelackofrequirementofMdj1pinthemt-Hsp70-mediatedproteinimportintomitochondria. Ontheotherhand,precursorproteinsintransitwhichhadfurtherenteredthematrixwerefoundinacomplex with Mdj1p. Our results suggest that Mdj1p together with mt-Hsp70 plays an important role as a chaperone formitochondriallysynthesizedpolypeptidechainsemergingfromtheribosomeandfortranslocatingproteins at a late import step.