Studies on fraction i protein. II. Comparison of physical, chemical, immunological and enzymatic properties between spinach and tobacco fraction i proteins

Kawashima, N.; Wildman, S.G.

Biochimica et Biophysica Acta 229(3): 749-760

1971


ISSN/ISBN: 0006-3002
PMID: 4995313
DOI: 10.1016/0005-2795(71)90293-5
Accession: 044428485

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Abstract
Fraction I proteins from tobacco and spinach were homogeneous and displayed Svedberg constants (s20,w) of 18.1 ± 0.1 and 18.2 ± 0.1, respectively. Mixing the two proteins produced neither separation nor skewness in the velocity sedimentation schlieren patterns. Both sodium dodecyl sulfate and alkali caused dissociation of the two proteins into two subunits which were readily separated from each other by Sephadex column chromatography. The ratio in amount of large to small subunits was the same for both tobacco and spinach Fraction I proteins. Disruption of SS bonds was not a requirement for dissociation and separation of subunits. Immunological analyses using antibodies against Fraction I protein from tobacco or spinach showed both proteins to be entirely homogeneous antigens, although the two proteins shared cross-reactive sites. Tryptic peptide analysis showed that 17 out of 22 (spinach) or 24 (tobacco) peptides of the large subunit were alike. In marked contrast, 9 out of 15 (spinach) or 10 out of 16 (tobacco) tryptic peptides of the small subunit were distinctly different. As for ribulose diphosphate carboxylase (Ec 4.1.1.39) activity, the Km for CO2 was the same for both tobacco and spinach Fraction I protein (1.7 · 10−2 M) whereas a small difference in Km for ribulose diphosphate was found (1.6 · 10−4 M for tobacco; 3.0 · 10−4 M for spinach).