CSM News Electronic Edition Volume 7, number 8 September 21, 1996 Please submit abstracts of your papers as soon as they have been accepted for publication by sending them to CSM-News@worms.cmb.nwu.edu. Back issues of CSM-News, the CSM Reference database and other useful information is available by anonymous ftp from worms.cmb.nwu.edu [165.124.233.50], via Gopher at the same address, or by World Wide Web at the URL "http://worms.cmb.nwu.edu/dicty.html" =========== Abstracts =========== MYOSIN MOTORS WITH ARTIFICIAL LEVER ARMS Michael Anson, Michael A. Geeves, Susanne E. Kurzawa and Dietmar J. Manstein National Institute for Medical Research, Mill Hill, London NW7 1AA, UK, Max-Planck-Institut fur Molekulare Physiologie, Postfach 102664, D-44026 Dortmund, Germany and Max-Planck-Institut fur Medizinische Forschung, Jahnstr. 29, D-69120 Heidelberg, Germany EMBO J., in press. Summary The myosin head consists of a globular catalytic domain and a light chain binding domain (LCBD). The coupling efficiency between adenosine triphosphate-hydrolysis and myosin-induced actin movement is known to decline as the LCBD is truncated or destabilized. However, it was not clear whether the observed alteration in the production of force and movement reflects only the mechanical changes to the length of the LCBD or whether these changes also affect the kinetic properties of the catalytic domain. Here we show that replacement of the LCBD with genetically engineered domains of similar rigidity and dimensions produces functional molecular motors with unchanged kinetic properties. The resulting single-chain, single-headed motors were produced in Dictyostelium discoideum and obtained after purification from a standard, peptone based growth medium at levels of up to 12 mg/l. Their actin-motility properties are similar or greater than those of native myosin. Rates of 2.5 mm s-1 and 3.3 mm s-1 were observed for motor domains fused to one or two of these domains, respectively. Their kinetic and functional similarity to the extensively studied myosin subfragment 1 (S1) and their accessibility to molecular genetic approaches makes these simple constructs ideal models for the investigation of chemo-mechanical coupling in the myosin motor. --------------------------------------------------------------------- DGAP1, a homoloque of rasGTPase activating proteins that controls growth, cytokinesis, and development in Dictyostelium discoideum Jan Faix and Werner Dittrich FEBS Lett., in press Abstract A protein accumulated in the cortical region of Dictyostelium discoideum cells proved to be a homologue of GTPase activating proteins, that are responsible for the inactivation of ras in yeast and man. Elimination of this protein, DGAP1, by gene replacement resulted in an increased rate of growth of D. discoideum cells on bacterial lawns, and in the formation of aberrant, multi-tipped fruiting bodies. Overexpression of DGAP1 caused the cells to become multi-nucleated since chromosome segregation during mitosis was not reliably followed by cleavage of the cells. These results suggest that in D. discoideum, ras or a related small GTP-binding protein is involved in regulating growth based on the phagocytosis of bacteria, and in coupling activities of the cell cortex to the organization of spindle and asters in mitotic cells. --------------------------------------------------------------------- Dictyostelium cytosolic fucosyltransferase synthesizes H type I trisaccharide in vitro M. Trinchera and S. Bozzaro Dept. Clinical and Biological Sciences, University of Turin, Italy FEBS Lett., in press. Abstract A fucosyltransferase activity is detected using lacto-N-biose as acceptor in the lower eukaryote Dictyostelium discoideum. Such transferase requires divalent cations and is inhibited by N-ethylmaleimideand detergent treatment. Apparent calculated Km values for GDP-Fuc and lacto-N-biose I are 1.27 mM and 2.80 mM, respectively. The activity is quantitatively recovered in the supernatant after centrifugation at 100,000 x g for 1 h. The reaction product, as determined by gel permeation chromatography, sensitivity to fucosidases, and analysis of partially methylated derivatives, is Fuca1-2Galb1-3GlcNAc (H type I trisaccharide) --------------------------------------------------------------------- [End CSM-News, volume 7, number 8]