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42. Annexin 24 from Capsicum annuum. X-ray structure and biochemical characterization.

Author

Hofmann, A, Proust, J, Dorowski, A, Schantz, R, and Huber, R

Abstract

This work provides the first three-dimensional structure of a member of the plant annexin family and correlates these findings with biochemical properties of this protein. Annexin 24(Ca32) from Capsicum annuum was purified as a native protein from bell pepper and was also prepared by recombinant techniques. To overcome the problem of precipitation of the recombinant wild-type protein in crystallization trials, two mutants were designed. Whereas an N-terminal truncation mutant turned out to be an unstable protein, the N-terminal His-tagged annexin 24(Ca32) was crystallized, and the three-dimensional structure was determined by x-ray diffraction at 2. 8 A resolution. The structure refined to an R-factor of 0.216 adopts the typical annexin fold; the detailed structure, however, is different from non-plant annexins, especially in domains I and III and in the membrane binding loops on the convex side. Within the unit cell there are two molecules per asymmetric unit, which differ in conformation of the IAB-loop. Both conformers show Trp-35 on the surface. The loop-out conformation is stabilized by tight interactions of this tryptophan with residue side chains of a symmetry-related molecule and enforced by a bound sulfate. Characterization of this plant annexin using biophysical methods revealed calcium-dependent binding to phospholipid vesicles with preference for phosphatidylcholine over phosphatidylserine and magnesium-dependent phosphodiesterase activity in vitro as shown with adenosine triphosphate as the substrate. A comparative unfolding study of recombinant annexin 24(Ca32) wild type and of the His-tag fusion protein indicates higher stability of the latter. The effect of this N-terminal modification is also visible from CD spectra. Both proteins were subjected to a FURA-2-based calcium influx assay, which gave high influx rates for the wild-type but greatly reduced influx rates for the fusion protein. We therefore conclude that the N-terminal domain is indeed a major regulatory element modulating different annexin properties by allosteric mechanisms.

Published
2000

48. Distributed Computation and TENEX-Related Activities

Author

BOLT BERANEK AND NEWMAN INC CAMBRIDGE MA, Schantz, R, Thomas, R, BOLT BERANEK AND NEWMAN INC CAMBRIDGE MA, Schantz, R, and Thomas, R

Abstract

This report describes efforts in the design of the National Software Works system and efforts to integrate TENEX into the National Software Works system.

Published
1977

49. Distributed Computation and TENEX-Related Activities

Author

BOLT BERANEK AND NEWMAN INC CAMBRIDGE MA, Schantz, R, Thomas, R, BOLT BERANEK AND NEWMAN INC CAMBRIDGE MA, Schantz, R, and Thomas, R

Abstract

This report describes efforts in the design of the National Software Works system and efforts to integrate TENEX into the National Software Works system., Sponsored in part by the DARPA.

Published
1976

50. CRONUS, A Distributed Operating System. Volume 4. CRONUS DOS implementation

Author

BBN LABS INC CAMBRIDGE MA, Schantz, R., Schroder, E., Barrow, M., Bono, G., Dean, M., Gurwitz, R., Lam, K., Lebowitz, K., Lipson, S., Neves, P., Sands, R., BBN LABS INC CAMBRIDGE MA, Schantz, R., Schroder, E., Barrow, M., Bono, G., Dean, M., Gurwitz, R., Lam, K., Lebowitz, K., Lipson, S., Neves, P., and Sands, R.

Abstract

This is the final report for the second contract phase for development of the Cronus Project. Cronus is the name given to the distributed operating system (DOS) and system architecture for distributed application development environment being designed and implemented by BBN Laboratories for the Air Force Rome Air Development Center (RADC). The project was begun in 1981. The Cronus distributed operating system is intended to promote resource sharing among interconnected computer systems and manage the collection of resources which are shared. Its major purpose is to provide a coherent and integrated system based on clusters of interconnected heterogeneous computers to support the development and use of distributed applications. Distributed applications range from simple programs that merely require convenient reference to remote data, to collections of complex distributions of Cronus is a unifying architecture and model for developing these distributed applications, as well as support for a number of system-provided functions which are common to many applications., See also Volume 1, ADA199986.

Published
1988

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