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Chimeric Arabidopsis thaliana
Ribulose-1,5-Bisphosphate Carboxylase/Oxygenase Containing a Pea Small
Subunit Protein Is Compromised in Carbamylation1
Timothy P. Getzoff,
Genhai Zhu,
Hans J. Bohnert*, and
Richard G. Jensen
Departments of Molecular and Cellular Biology (T.P.G., H.J.B.),
Biochemistry (G.Z., H.J.B., R.G.J.), and Plant Sciences (H.J.B.,
R.G.J.), The University of Arizona, Tucson, Arizona 85721-0088
A cDNA of pea (Pisum
sativum L.) RbcS 3A, encoding a small subunit
protein (S) of ribulose-1,5-bisphosphate carboxylase/oxygenase (Rubisco), has been expressed in Arabidopsis thaliana
under control of the cauliflower mosaic virus 35S promoter, and the
transcript and mature S protein were detected. Specific antibodies
revealed two protein spots for the four Arabidopsis S and one
additional spot for pea S. Pea S in chimeric Rubisco amounted to 15 to
18% of all S, as judged by separation on two-dimensional isoelectric focusing/sodium dodecyl sulfate-polyacrylamide gel electrophoresis gels
from partially purified enzyme preparations and quantitation of
silver-stained protein spots. The chimeric enzyme had 11 ± 1%
fewer carbamylated sites and a 11 ± 1% lower carboxylase
activity than wild-type Arabidopsis Rubisco. Whereas pea S expression, preprotein transport, and processing and assembly resulted in a stable
holoenzyme, the chimeric enzyme was reproducibly catalytically less
efficient. We suggest that the presence of, on average, one foreign S
per holoenzyme is responsible for the altered activity. In addition,
higher-plant Rubisco, unlike the cyanobacterial enzyme, seems to have evolved species-specific interactions between S and the
large subunit protein that are involved in carbamylation of the active
site.
1
The work was supported by the National Science
Foundation (Biochemistry Program, 1991-1994), by Japan Tobacco, and in
part by the Arizona Agricultural Experiment Station.
*
Corresponding author; e-mail bohnerth{at}u.arizona.edu; fax
1-520-621-1697.
Plant Physiol. (1998) 116: 695-702
Copyright Clearance Center: 0032-0889/98/116/0695/08
© 1998 American Society of Plant Physiologists
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