An essential role for water in an enzyme reaction mechanism: the crystal structure of the thymidylate synthase mutant E58Q

A water-mediated hydrogen bond network coordinated by glutamate 60(58) appears to play an important role in the thymidylate synthase (TS) reaction mechanism. We have addressed the role of glutamate 60(58) in the TS reaction by cocrystalizing the Escherichia coli TS mutant E60(58)Q with dUMP and the cofactor analog CB3717 and have determined the X-ray crystal structure to 2.5 A resolution with a final R factor of 15.2% (Rfree = 24.0%). Using difference Fourier analysis, we analyzed directly the changes that occur between wild-type and mutant structures. The structure of the mutant enzyme suggests that E60(58) is not required to properly position the ligands in the active site and that the coordinated hydrogen bond network has been disrupted in the mutant, providing an atomic resolution explanation for the impairment of the TS reaction by the E60(58)Q mutant and confirming the proposal that E60(58) coordinates this conserved hydrogen bond network. The structure also provides insight into the role of specific waters in the active site which have been suggested to be important in the TS reaction. Finally, the structure shows a unique conformation for the cofactor analog, CB3717, which has implications for structure-based drug design and sheds light on the controversy surrounding the previously observed enzymatic nonidentity between the chemically identical monomers of the TS dimer.     

The critical care of the severely injured patient--II. Pulmonary and renal support

The management of pulmonary support in trauma patients is a balance of risk versus benefit. Decisions must be based upon the patient's physiologic needs, with constant attention to the potential complications of the therapy. The avoidance of renal failure by volume replacement, careful attention to the use of potentially nephrotoxic agents, and close monitoring of renal function can reduce the incidence and severity of this serious complication of trauma. If renal failure does occur, aggressive renal replacement therapy with either intermittent hemodialysis or CRRT should be tailored to the clinical situation.     

The characteristics of gastroduodenal pathology in the workers of a plant for plastic construction materials

Out of 162 workers exposed to chemicals in the manufacture of construction plastics an outpatient and inpatient examination revealed gastrointestinal problems in 84.3%, vegetative nervous disorders in 62.6% of the examinees. In a random sample of 47 workers high acid production occurred in 63.3% and erosions, ulcers, cicatrices in the stomach and duodenum in 45.2% of them. No HLA genetic predisposition to duodenal ulcer was reported. Histological findings on gastroduodenal mucosa biopsies were indicative of the dystrophy in the chief and lining cells of the gastric glands, epithelium and stroma with formation of the dark cells. The authors insist on consideration of the above facts in examination and treatment of workers exposed to chemicals at construction plastics plants.     

Immunogenicity of synthetic HIV-1 gp120 V3-loop peptide-conjugate immunogens

A successful prophylactic human immunodeficiency virus type 1 (HIV-1) vaccine must elicit an immune response that will prevent establishment of the persistent viral infection. The only response shown to be effective in this regard is virus-neutralizing antibody directed against the viral gp120 hypervariable V3-loop region. Conjugate immunogens, containing cyclic peptides representing the V3 determinant covalently bound to a carrier protein, were capable of eliciting virus-neutralizing antibodies. The consistency of the response was related to peptide size. The smaller cyclic peptides, expressing relatively conserved sequences from the V3-loop apex, were poor inducers of neutralizing activity. In contrast, the largest cyclic peptides mediated neutralizing responses that were similar to those observed and previously reported for intact gp120 immunogens. A cyclic synthetic peptide expressing most of the prototypic HIV-1 MN variant V3 determinant warrants further study as a potentially effective vaccine immunogen.     

Evoked potentials in the dog sensomotor cortex during an avoidance reaction

Elaboration of avoidance reaction (conditioned reflex of the second order), based on a preliminarily elaborated escape (conditioned reflex of the first order) from pulse electro-cutaneus stimulation of the paw, led to a well-defined reorganization of the activity of the cortical limb projection structures. By excitability parameters (EP and motor reaction thresholds) and by changes of parameters of EP to electro-cutaneous stimulation, in escape reactions the sensory functions are drastically enhanced. Judging from the EP pattern, in response to the first pulses of the series (intensification of late components, appearance of an additional late negative wave), these pulses act as signals. With the introduction of distant stimuli (clicks), the electric current loses its signal significance, and the EP produced by it are sharply depressed and simplified. In the somatosensory projection of the paw, EP to clicks observed before conditioning, decrease or disappear completely. But they appear in the motor projection of the paw. In conjunction with excitation parameters, these data suggest the predominance of efferent functions of the paw in avoidance reactions.     

Carotid body hypertrophy in patients with cystic fibrosis and cyanotic congenital heart disease

Austrobilharzia terrigalensis from Egretta sacra and Larus novaehollandiae at Heron Island, Great Barrier Reef, is described. The prosobranch snail Planaxis sulcatus is the natural intermediate host of A. terrigalensis at Heron Island. The life-cycle of A. terrigalensis was worked out using naturally and experimentally infected Planaxis sulcatus and experimentally infected chickens. Experiments to produce a dermatitis with cercariae of A. terrigalensis were negative. Austrobilharzia terrigalensis infects a wide range of natural hosts in Canada, continental United States, Hawaii and Australia, and uses different snail genera belonging to different families as intermediate hosts in different areas. The wide distribution is apparently due to the habits of some hosts which migrate between the northern and southern hemispheres.