The prominent 28 kDa polypeptide in clathrin coated vesicle fractions from developing pea cotyledons is contaminating ferritin |
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| Authors: | Birgit Hoh David G. Robinson |
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| Affiliation: | 1. Goethe University, Department of Biosciences, Molecular Cell Biology of Plants, Frankfurt/Main, Germany;2. SPOT-ITN Consortium;3. University of Vienna, Department of Ecogenomics and Systems biology, Vienna, Austria;4. School of Biotechnology and Bioinformatics, D.Y. Patil University, Mumbai, India;5. Goethe University, Cluster of Excellence Frankfurt, Frankfurt/Main, Germany;6. Goethe University, Buchmann Institute of Molecular Life Sciences, Frankfurt/Main, Germany;1. National Laboratory of Biomacromolecules, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China;2. Key Laboratory of RNA Biology, CAS Center for Excellence in Biomacromolecules, Institute of Biophysics, Chinese Academy of Sciences, 15 Datun Road, Beijing 100101, China;3. College of Life Sciences, University of Chinese Academy of Sciences, Beijing 100049, China |
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| Abstract: | SDS-PAGE of clathrin coated vesicle (CCV) fractions prepared from developing pea cotyledons are characterized by the presence of a 28 kDa polypeptide. Like clathrin light chains, this polypeptide is heat-stable and can bind calcium ions. However, the distribution of this polypeptide is not identical to that of clathrin in the rate zonal gradients used to purify CCV. Negatively stained preparations reveal small, 12 nm diameter hollow particles, in addition to CCV. As judged by the electron dense centre to these particles, we infer that CCV preparations from pea cotyledons are contaminated with phytoferritin. This has been confirmed by immuno-blotting with pea ferritin antibodies. The degree of phytoferritin contamination in CCV fractions is greater when RNase digestion of postmicrosomal pellets is performed at cold, rather than warm temperatures. |
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