Localization and properties of kinases in clathrin-coated vesicles from zucchini hypocotyls

被引:3
作者
Drucker, M [1 ]
Happel, N [1 ]
Robinson, DG [1 ]
机构
[1] UNIV GOTTINGEN,INST PFLANZENPHYSIOL,D-37073 GOTTINGEN,GERMANY
来源
EUROPEAN JOURNAL OF BIOCHEMISTRY | 1996年 / 240卷 / 03期
关键词
adaptins; clathrin-coated vesicles; kinase; uncoating adenosinetriphosphatase; zucchini;
D O I
10.1111/j.1432-1033.1996.0570h.x
中图分类号
Q5 [生物化学]; Q7 [分子生物学];
学科分类号
071010 ; 081704 ;
摘要
Five major polypeptides of 70, 50, 47, 19 and 17 kDa and four minor polypeptides (100, 65, 45 and 39 kDa) become phosphorylated when clathrin-coated vesicles (CCV) from zucchini hypocotyls are incubated in [gamma(32)P]Mg-ATP. After dissociation with 0.5 M Tris/HCl the CCV coat polypeptides were subjected to gel filtration in order to separate clathrin triskelions from beta-adaptin-containing fractions. Only the latter bore kinase activities, with phosphorylated polypeptides of 39 kBa in addition to the 50, 19-kDa and 17-kDa polypeptides just mentioned, Heparin, an inhibitor of casein kinase II, permitted the phosphorylation of only the 19-kDa and 17-kDa polypeptides. Staurosporine, an inhibitor of protein kinase c-like acitivities, prevented the phosporylation of the 70-kDa polypeptide. When recombined with the triskelions the beta-adaptin fractions achieved the phosphorylation of the 45-kDa and 70-kDa polypeptides. Because of its heat stability and calcium-binding properties we interpret the 45-kDa polypeptide as being a clathrin light chain. Antibodies raised against the 70-kDa group of heat-shock proteins (Hsp70) recognize a 70-kDa polypeptide in the beta-adaptin-containing fractions. Because this polypeptide only phosphorylates in the presence of triskelions we consider it to be the uncoating ATPase, which is known to aggregate upon dissociation of the CCV coat. Our results therefore indicate that zucchini CCV contain a number of phosphorylable polypeptides equivalent to the beta, mu and sigma adaptins of bovine brain. Just as in bovine brain CCV a casein-kinase-II-like activity is associated with the zucchini CCV 50/47-kDa polypeptides, further pointing to their identity as plant mu(2)/mu(1) adaptin equivalents.
引用
收藏
页码:570 / 575
页数:6
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