Allosteric loss-of-function mutations in HIV-1 Nef from a long-term non-progressor

Activation of Src family kinases by human immunodeficiency virus type 1 (HIV-1) Nef may play an important role in the pathogenesis of HIV/AIDS. Here we investigated whether diverse Nef sequences universally activate Hck, a Src family member expressed in macrophages and other HIV-1 target cells. In general, we observed that Hck activation is a highly conserved Nef function. However, we identified an unusual Nef variant from an HIV-positive individual that did not develop AIDS which failed to activate Hck despite the presence of conserved residues linked to Hck SH3 domain binding and kinase activation. Amino acid sequence alignment with active Nef proteins revealed differences in regions not previously implicated in Hck activation, including a large internal flexible loop absent from available Nef structures. Substitution of these residues in active Nef compromised Hck activation without affecting SH3 domain binding. These findings show that residues at a distance from the SH3 domain binding site influence Nef interactions allosterically with a key effector protein linked to AIDS progression.     

Biotransformation Patterns of 2,4,6-Trinitrotoluene by Aerobic Bacteria

2,4,6-Trinitrotoluene (TNT), a toxic nitroaromatic explosive, accumulates in the environment, making necessary the remediation of contaminated areas and unused materials. Although bioremediation has been utilized to detoxify TNT, the metabolic processes involved in the metabolism of TNT have proven to be complex. The three aerobic bacterial strains reported here (Pseudomonas aeruginosa, Bacillus sp., and Staphylococcus sp.) differ in their ability to biotransform TNT and in their growth characteristics in the presence of TNT. In addition, enzymatic activities have been identified that differ in the reduction of nitro groups, cofactor preferences, and the ability to eliminate-NO₂ from the ring. The Bacillus sp. has the most diverse bioremediation potential owing to its growth in the presence of TNT, high level of reductive ability, and capability of removing-NO₂ from the nitroaromatic ring. Received: 16 May 1997 / Accepted: 19 July 1997     

Mandibular movements and their control inHomarus gammarus

Summary The skeletal morphology, musculature and innervation of the mandible of the common lobster,Homarus gammarus, are described as a basis for the functional study included in the two subsequent papers.Although the mandible articulation takes the form of a hinge with movement in a single plane, the musculature of the mandible is complex. The main muscles are similar to those ofAstacus (Schmidt, 1915) but some smaller, previously undescribed muscles were found.As forAstacus (Keim, 1915) andCambarus (Chaudonneret, 1956) the mandibular muscles are innervated by two nerve trunks, the inner and outer mandibular nerves. However, differences occur in the branching of these nerves and the muscles which they innervate.A group of sensory cells associated with the posterior stomach nerve (omn 4) are described. It is suggested that these form a proprioceptive organ associated with the hypodermis overlying the lateral mandible articulation.An interesting group of neurones lying at the confluence of nerve branches from omn 2, omn 3, and omn 4 is described.     

Energy landscapes and properties of biomolecules

Thermodynamic and dynamic properties of biomolecules can be calculated using a coarse-grained approach based upon sampling stationary points of the underlying potential energy surface. The superposition approximation provides an overall partition function as a sum of contributions from the local minima, and hence functions such as internal energy, entropy, free energy and the heat capacity. To obtain rates we must also sample transition states that link the local minima, and the discrete path sampling method provides a systematic means to achieve this goal. A coarse-grained picture is also helpful in locating the global minimum using the basin-hopping approach. Here we can exploit a fictitious dynamics between the basins of attraction of local minima, since the objective is to find the lowest minimum, rather than to reproduce the thermodynamics or dynamics.     

Disordered development of macrophages in non-Hodgkin’s lymphoma

Macrophage development in 20 untreated patients with non-Hodgkin’s lymphoma (NHL) has been studied and compared with that in 20 normal subjects. Morphometric measurements were carried out on ultrastructural features of cell, nucleus and mitochondria during 6 days suspension culture of blood monocytes in the presence of autologous serum and lymphocytes. The results were subjected to multivariate and univariate analysis of variance. Statistically significant differences were found between the subject groups with respect to the volumes and surface areas of cell, nucleus and mitochondria, to the excess surface membrane of cell and nucleus (as compared with equivalent spheres) and to the number of mitochondrial profiles per section. It would appear that the patients’ cell grew less, showed less elaboration of surface features and had reduced nuclear and mitochondrial development, the latter affecting mitochondrial numbers rather than individual size. The findings provide further evidence that mononuclear phagocytes are deranged in NHL.     

Some foundations of contemporary methods for culturing preimplantation embryos; a personal account

This article traces the history of the work on mammalian embryos carried out by the author, his students and related scientists. It traces the work from the initial experiments at the King Ranch Laboratories, University of Pennsylvania in 1966, the set up of an embryo culture laboratory at the Department of Veterinary Physiology, University of Sydney and the work within that laboratory from 1967 to 1974. This is followed by an account of the author's subsequent work at Murdoch University from 1975 till his retirement in 1996. The significant role of some of the author's graduate students in human IVF is also documented.     

Hydrogen deuterium exchange mass spectrometry applied to chaperones and chaperone-assisted protein folding

Introduction: Hydrogen–deuterium exchange (HDX) mass spectrometry (MS) is ideal for monitoring the protein folding and unfolding. The exchange of a deuterium in solution for an amide hydrogen in a protein can be very different depending on the degree of folding and protection of backbone amide positions. Molecular chaperones that assist with protein folding in vivo are necessary for folding of many substrate (client) proteins. HDX MS provides valuable insight into what chaperones are doing in protein folding and how they are doing it.

Areas covered: Application of HDX MS to the protein folding problem was desirable from the outset of the technique, but technical issues prohibited many studies. In the last 20 years, conformational changes of chaperones themselves (e.g., GroEL/GroES, Hsp70, and Hsp90) have been studied. Studies of interactions between chaperones, co-chaperones, and substrate proteins have revealed binding interfaces, allosteric conformational changes, and remodeling of components during various chaperone cycles. Experiments elucidating how chaperones contribute to and enhance the folding pathway of substrate proteins have been demonstrated.

Expert opinion: Technical issues that once prevented the analysis of chaperones have largely been resolved, permitting exciting comprehensive HDX MS studies of folding pathways during chaperone-assisted protein folding.