title: Subunit Asa1 spans all the peripheral stalk of the mitochondrial ATP synthase of the chlorophycean alga Polytomella sp.
abstract: Mitochondrial F1FO-ATP synthase of chlorophycean algae is dimeric. It contains eight orthodox subunits (alpha, beta, gamma, delta, epsilon, OSCP, a and c) and nine atypical subunits (Asa1 to 9). These subunits build the peripheral stalk of the enzyme and stabilize its dimeric structure. The location of the 66.1kDa subunit Asa1 has been debated. On one hand, it was found in a transient subcomplex that contained membrane-bound subunits Asa1/Asa3/Asa5/Asa8/a (Atp6)/c (Atp9). On the other hand, Asa1 was proposed to form the bulky structure of the peripheral stalk that contacts the OSCP subunit in the F1 sector. Here, we overexpressed and purified the recombinant proteins Asa1 and OSCP and explored their interactions in vitro, using immunochemical techniques and affinity chromatography. Asa1 and OSCP interact strongly, and the carboxy-terminal half of OSCP seems to be instrumental for this association. In addition, the algal ATP synthase was partially dissociated at relatively high detergent concentrations, and an Asa1/Asa3/Asa5/Asa8/a/c10 subcomplex was identified. Furthermore, Far-Western analysis suggests an Asa1-Asa8 interaction. Based on these results, a model is proposed in which Asa1 spans the whole peripheral arm of the enzyme, from a region close to the matrix-exposed side of the mitochondrial inner membrane to the F1 region where OSCP is located. 3D models show elongated, helix-rich structures for chlorophycean Asa1 subunits. Asa1 subunit probably plays a scaffolding role in the peripheral stalk analogous to the one of subunit b in orthodox mitochondrial enzymes.
authors.affiliation: Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, México D.F., Mexico.
authors.forename: Lilia
authors.initials: L
authors.lastname: Colina-Tenorio
authors.affiliation: Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, México D.F., Mexico.
authors.forename: Héctor
authors.initials: H
authors.lastname: Miranda-Astudillo
authors.affiliation: Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, México D.F., Mexico.
authors.forename: Araceli
authors.initials: A
authors.lastname: Cano-Estrada
authors.affiliation: Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, México D.F., Mexico.
authors.forename: Miriam
authors.initials: M
authors.lastname: Vázquez-Acevedo
authors.affiliation: Genetics and Physiology of Microalgae, Department of Life Sciences, University of Liège, B-4000, Liège, Belgium.
authors.forename: Pierre
authors.initials: P
authors.lastname: Cardol
authors.affiliation: Genetics and Physiology of Microalgae, Department of Life Sciences, University of Liège, B-4000, Liège, Belgium.
authors.forename: Claire
authors.initials: C
authors.lastname: Remacle
authors.affiliation: Instituto de Fisiología Celular, Universidad Nacional Autónoma de México, México D.F., Mexico. Electronic address: dhalphen@ifc.unam.mx.
authors.forename: Diego
authors.initials: D
authors.lastname: González-Halphen
mesh_terms: D000595:Amino Acid Sequence; D000460:Chlorophyta / Q000201:enzymology*; D025261:Mitochondrial Proton-Translocating ATPases / Q000737:chemistry*; D008969:Molecular Sequence Data; D021122:Protein Subunits
chemical_list: D021122:Protein Subunits; D025261:Mitochondrial Proton-Translocating ATPases
keywords: Asa subunits; Chlamydomonas reinhardtii; Chlorophycean algae; Dimeric mitochondrial complex V; F(1)F(O)-ATP synthase peripheral stalk; Polytomella sp.