The H$ sp+$-translocating inorganic pyrophosphatase from vacuolar membranes of red beet storage roots (Beta vulgaris L.) was purified after solubilization in Triton X-100 through a combination of anion-exchange and size exclusion chromatographies. SDS-PAGE showed strong correlation between a 67 kDa polypeptide and pyrophosphatase activity. Radiation-inactivation studies of the H$ sp+$-PPase indicate a functional size of 91 kDa for hydrolysis and 320 kDa for H$ sp+$ translocation, suggesting an oligomeric structure for the holoenzyme. Affinity purified antibodies were used to screen cDNA libraries of Arabidopsis thaliana yielding clones which contained sequences matching amino acid sequences obtained from tryptic fragments of the 67 kDa hydrolytic subunit. The predicted protein is highly hydrophobic with a molecular size of 81 kDa. Southern analyses show a single copy for the H$ sp+$-PPase in Arabidopsis. The lack of sequence identities between the H$ sp+$-PPase and other known proteins implies a novel class of ion translocases.
| Identifer | oai:union.ndltd.org:LACETR/oai:collectionscanada.gc.ca:QMM.39460 |
| Date | January 1992 |
| Creators | Sarafian, Vahé |
| Publisher | McGill University |
| Source Sets | Library and Archives Canada ETDs Repository / Centre d'archives des thèses électroniques de Bibliothèque et Archives Canada |
| Language | English |
| Detected Language | English |
| Type | Electronic Thesis or Dissertation |
| Format | application/pdf |
| Coverage | Doctor of Philosophy (Department of Biology.) |
| Rights | All items in eScholarship@McGill are protected by copyright with all rights reserved unless otherwise indicated. |
| Relation | alephsysno: 001317720, proquestno: NN80465, Theses scanned by UMI/ProQuest. |
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