期刊
PHOTOSYNTHESIS RESEARCH
卷 118, 期 3, 页码 209-218出版社
SPRINGER
DOI: 10.1007/s11120-013-9916-0
关键词
CO2/O-2 specificity; Directed mutagenesis; Photosynthesis; Protein structure; Rubisco; Rubisco activase
资金
- Division of Chemical Sciences, Geosciences, and Biosciences, Office of Basic Energy Sciences, of the United States Department of Energy [DE-FG02-00ER15044, DE-AI02-97ER20268]
- U.S. Department of Energy (DOE) [DE-FG02-00ER15044] Funding Source: U.S. Department of Energy (DOE)
Ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco) catalyzes the initial step of carbon metabolism in photosynthesis. The holoenzyme comprises eight large subunits, arranged as a tetramer of dimers around a central solvent channel that defines a fourfold axis of symmetry, and eight small subunits, arranged as two tetramers at the poles of the axis. The phylogenetically divergent small-subunit loops between beta-strands A and B form the entrance to the solvent channel. In the green alga Chlamydomonas reinhardtii, Ile-58 from each of the four small-subunit beta A-beta B loops defines the minimal diameter of the channel opening. To understand the role of the central solvent channel in Rubisco function, directed mutagenesis and transformation of Chlamydomonas were employed to replace Ile-58 with Ala, Lys, Glu, Trp, or three Trp residues (I58W3) to close the entrance to the channel. The I58E, I58K, and I58W substitutions caused only small decreases in photosynthetic growth at 25 and 35 A degrees C, whereas I58W3 had a substantial effect at both temperatures. The mutant enzymes had decreased carboxylation rates, but the I58W3 enzyme had decreases in both carboxylation and CO2/O-2 specificity. The I58E, I58W, and I58W3 enzymes were inactivated at lower temperatures than wild-type Rubisco, and were degraded at slower rates under oxidative stress. However, these mutant enzymes were activated by Rubisco activase at normal rates, indicating that the structural transition required for carboxylation is not affected by altering the solvent channel opening. Structural dynamics alone may not be responsible for these distant effects on the Rubisco active site.
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