This Article Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Services Email this article to a friend Similar articles in this journal Similar articles in Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via HighWire Citing Articles via CrossRef Citing Articles via Web of Science (29) Citing Articles via Google Scholar Google Scholar Articles by Roesler, K. R. Articles by Ogren, W. L. Search for Related Content PubMed PubMed Citation Articles by Roesler, K. R. Articles by Ogren, W. L. Agricola Articles by Roesler, K. R. Articles by Ogren, W. L.

Metabolism and Enzymology

Chlamydomonas reinhardtii Phosphoribulokinase ¹

Sequence, Purification, and Kinetics

Department of Agronomy, University of Illinois at Urbana-Champaign, Urbana, Illinois 61801, U.S. Department of Agriculture, Agricultural Research Service, Urbana, Illinois 61801

The sequence and kinetic properties of phosphoribulokinase purified from Chlamydomonas reinhardtii were determined and compared with the spinach (Spinacea oleracea) enzyme. Chlamydomonas phosphoribulokinase was purified to apparent homogeneity, with a specific activity of 410 micromoles per minute per milligram. Polyclonal antibodies to the purified protein were used to isolate a Chlamydomonas cDNA clone, which, upon sequencing, was found to contain the entire coding region. The transit peptide cleavage site was determined by Edman analysis of the mature protein. The precursor protein consists of a 31 amino acid transit peptide and a 344 amino acid mature polypeptide. The mature polypeptide has a calculated molecular weight of 38.5 kilodaltons and a pl of 5.75. The V_max of the purified enzyme was 465 micromoles per minute per milligram, with apparent K_m values of 62 micromolar ATP and 56 micromolar ribulose 5-phosphate. Immunoblot analysis indicated antigenic similarity and a similar subunit size for the enzyme from five higher plant species and Chlamydomonas. Southern blot analysis of Chlamydomonas genomic DNA indicated the presence of a single phosphoribulokinase gene. Comparison of the mature proteins from Chlamydomonas and spinach revealed 86 amino acid differences in primary structure (25% of the total) without a major difference in kinetic properties. The transit peptides of the spinach and Chlamydomonas proteins possessed little sequence homology.

This article has been cited by other articles:

				M. E. Rumpho, S. Pochareddy, J. M. Worful, E. J. Summer, D. Bhattacharya, K. N. Pelletreau, M. S. Tyler, J. Lee, J. R. Manhart, and K. M. Soule Molecular Characterization of the Calvin Cycle Enzyme Phosphoribulokinase in the Stramenopile Alga Vaucheria litorea and the Plastid Hosting Mollusc Elysia chlorotica Mol Plant, November 1, 2009; 2(6): 1384 - 1396. [Abstract] [Full Text] [PDF]

				D. Kobayashi, M. Tamoi, T. Iwaki, S. Shigeoka, and A. Wadano Molecular Characterization and Redox Regulation of Phosphoribulokinase from the Cyanobacterium Synechococcus sp. PCC 7942 Plant Cell Physiol., March 15, 2003; 44(3): 269 - 276. [Abstract] [Full Text] [PDF]

				S. Lebreton and B. Gontero Memory and Imprinting in Multienzyme Complexes. EVIDENCE FOR INFORMATION TRANSFER FROM GLYCERALDEHYDE-3-PHOSPHATE DEHYDROGENASE TO PHOSPHORIBULOKINASE UNDER REDUCED STATE IN CHLAMYDOMONAS REINHARDTII J. Biol. Chem., July 23, 1999; 274(30): 20879 - 20884. [Abstract] [Full Text] [PDF]

				T. Hariharan, P. J. Johnson, and R. Ann Cattolico Purification and Characterization of Phosphoribulokinase from the Marine Chromophytic Alga Heterosigma carterae Plant Physiology, May 1, 1998; 117(1): 321 - 329. [Abstract] [Full Text]