. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. 204 REPORTS FROM THE MBL GENERAL SCIENTIFIC MEETINGS Dr. S. Brady) to determine whether kinesin binds to myosin V in squid neurons. In addition, we used recombinanl tail frag- ments of mouse MyoV and mouse kinesin (constructs provided by Dr. Huang) to test whether these fragments form heterolo- gous complexes with squid brain kinesin and myosin V, respec- tively. The plasmids containing the glutathione S-transferase (GST)-labeled mouse AF6/cno tail-globular-domain (GST- MyoV-tail aa!569 to aa!768 without the coiled medial ta
. The Biological bulletin. Biology; Zoology; Biology; Marine Biology. 204 REPORTS FROM THE MBL GENERAL SCIENTIFIC MEETINGS Dr. S. Brady) to determine whether kinesin binds to myosin V in squid neurons. In addition, we used recombinanl tail frag- ments of mouse MyoV and mouse kinesin (constructs provided by Dr. Huang) to test whether these fragments form heterolo- gous complexes with squid brain kinesin and myosin V, respec- tively. The plasmids containing the glutathione S-transferase (GST)-labeled mouse AF6/cno tail-globular-domain (GST- MyoV-tail aa!569 to aa!768 without the coiled medial tail domain) and the histidine (His)-labeled, kinesin-tail fragment of mouse ubiquitous kinesin (aa763-856) were expressed in E. cnli. The bacterially expressed mouse GST- or His-tagged frag- ments were purified on affinity columns and used in binding assays of squid brain extracts. The specificity of aQLLQ and H2 was determined by probing blots of squid brain extracts with the two antibodies. aQLLQ recognized a single protein of 196 kD in blots of squid brain extracts (Fig. 1A, lane 2), and H2 recognized a single protein of 120 kD (Fig. 1A. lane 3). These antibodies were then used in immunoprecipitation experiments to determine whether myosin V and kinesin interact directly. Squid brain extracts were incubated with the aQLLQ and then with protein-A beads. The precipitate was collected by centrifugation and run on a SDS-PAGE gel (Fig. IB, lane 1). Proteins in the 50-55 kD range representing the antibody were the primary components in the precipitate. A blot of the immuno-precipitate probed with aQLLQ showed a myosin-V band at 196 kD, protein bands at 50-55 kD representing the antibody, and one unknown protein at about 100 kD. The unknown protein may represent a serum protein that binds to the antibody Antibodies. 2 3 IP: QLLQ Ab: QLLQ H2 H2 H2 rf* B C Figure 1. Protein si:e murker is listed on the left, dashes indicate the corresponding vi;c\ on the gels In the right. Pane/ A: La
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Keywords: ., bookauthorlilliefrankrat, booksubjectbiology, booksubjectzoology
