Partial deficiency of hypoxanthine-guanine phosphoribosyltransferase with reduced affinity for PP-ribose-P in four related males with gout

Summary A family is described in which four affected males, spanning two generations, have hyperuricemia and gout accompanied by hematuria but are without severe neurologic involvement. The affected males were found to have markedly reduced levels of erythrocytic hypoxanthine-guanine phosphoribosyltransferase (HGPRT) activity; these were 5–12% with hypoxanthine and 0.5–3% with guanine as compared to controls. Erythrocytic adenine phosphoribosyltransferase (APRT) was approximately three-fold elevated in the affected individuals.The residual HGPRT activity in affected males enabled characterization of some of the properties of this mutation. The apparent Michaelis constants (km) for both hypoxanthine and guanine were essentially unchanged, whereas the km for PP-ribose-P was approximately 10–20-fold elevated for all four affected males. The enzyme was more sensitive to product inhibition by IMP and GMP than controls, and exhibited greater thermal lability at 65°C than found with control lysates.     

Thermal sensitivity of scope for activity in Pagothenia borchgrevinki, a cryopelagic Antarctic nototheniid fish

The thermal sensitivity of scope for activity was studied in the Antarctic nototheniid fish Pagothenia borchgrevinki. The scope for activity of P. borchgrevinki at 0°C was 189 mg O₂ kg⁻¹ h⁻¹ (factorial scope 6.8) which is similar to that of temperate and tropical species at their environmental temperatures, providing no evidence for metabolic cold adaptation of maximum activity. The scope for activity increased to a maximum value of 266 mg O₂ kg⁻¹ h⁻¹ (factorial scope 8.3) at 3°C and then decreased from 3 to 6°C. The thermal sensitivity of critical swimming speed was also investigated and followed a similar pattern to aerobic scope for activity, suggesting oxygen limitation of aerobic performance. Oxygen consumption rates and ventilation frequencies were monitored for 24 h after the swimming challenge and the recovery of both parameters to resting levels was rapid and independent of temperature.     

The catalytic site residues and interfacial binding of human pancreatic lipase.

In this study, the essential serine residue and 2 other amino acids in human pancreatic triglyceride lipase (triacylglycerol acylhydrolase, EC 3.1.1.3) were tested for their contribution to the enzyme's catalytic site or interfacial binding site. By site-specific mutagenesis of the cDNA for human pancreatic lipase, amino acid substitutions were made at Ser153, His264, and Asp177. The mutant cDNAs were expressed in transfected COS-1 cells. Both the medium and the cells were examined for the presence of pancreatic lipase by Western blot analysis. The activity of the expressed proteins against triolein and the interfacial binding was measured. Proteins with mutations in Ser153 were secreted by the cells and bound to interfaces but had no detectable activity. Changing His264 to a leucine or Asp177 to an asparagine also produced inactive lipase. Substituting glutamic acid for Asp177 produced an active protein. These results demonstrate that Ser153 is involved in the catalytic site of pancreatic lipase and is not crucial for interfacial binding. Moreover, the essential roles of His264 and Asp177 in catalysis were demonstrated. A Ser-His-Asp catalytic triad similar to that present in serine proteases is present in human pancreatic lipase.     

Fluorescence anisotropy from diphenylhexatriene in rat liver plasma membranes

Rat livers were fractionated to obtain intracellular membrane preparations and a highly purified preparation of bile canaliculi. The fraction containing bile canaliculi was homogenized and subfractionated to give fractions representing fragments of contiguous membrane and of canalicular microvilli. The relative purity and extent of contamination of each preparation was determined. When the fluorescent probe 1,6-diphenyl-1,3,5-hexatriene was incorporated into aliquots of each fraction at the same probe: lipid ratio and the steady-state anisotropy of its fluorescence measured, it was found that the plasma membrane preparations were much more ordered than the intracellular membrane preparations. Of the plasma membrane preparations, that containing the canalicular microvilli was the most ordered, even allowing for any contribution of contaminants. Thus the microvillus membrane of the bile canaliculus appears to be the most ordered domain of the plasma membrane of the hepatocyte. The high order in this domain may be a factor in reducing the susceptibility to bile salt damage during bile secretion, since it is this region which is exposed to high concentrations of bile salts in vivo.     

Evidence for histidine in the active site of papain

Papain was irreversibly inhibited by 1,3-dibromoacetone, a reagent designed to react first with the active-site cysteine residue and subsequently with a second nucleophile. The molecular weight of the inhibited enzyme was indistinguishable from that of papain itself, and no evidence of dimeric or oligomeric species was found. The optical-rotatory-dispersion curves of chloroacetone-inhibited papain and 1,3-dibromoacetone-inhibited papain were essentially similar. Amino acid analysis of the 1,3-dibromo[2-¹⁴C]acetone-inhibited enzyme and the performic acid-oxidized material clearly showed that a cysteine and histidine residue had been alkylated through the thiol and N-1 of the imidazole group respectively. These groups must therefore be within 5å of each other in the tertiary structure of papain. Possible mechanistic implications are briefly discussed.     

Interspecific differences in the biology of aphids (homoptera: Aphididae) on leaves of Vicia faba I. Feeding behaviour

Aphids of six species settled on the undersurface of bean leaves in characteristic patterns. Some species, e.g. Aphis fabae, settled wholly on veins. In others, e.g. Myzus spp. a considerable proportion settled away from veins, especially on the leaf margin. Microscopic studies showed that the probes of A. fabae were directed to the phloem in the veins as were those probes of Myzus spp. which entered the veins. Many probes of Myzus passed into spongy mesophyll alone. This apparently offered an alternative food source to them. The interspecific differences, which were matched by differences between age groups within species, are explained in terms of the differing proportions of the aphid populations exploiting the alternative food sources and of preferences for veins of a particular size.

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Zusammenfassung Blattläuse sechs verschiedener Arten, die sich ungehindert auf reifen Blättern von Vicia faba ansiedeln konnten, bildeten arttypische Verteilungen. Auf diese Weise wurde eine Übergangsreihe von Arteigentümlichkeiten deutlich zwischen Aphis fabae und Megoura viciae, die sich an den großen Adern ansiedelten, und Myzus ornatus, welche sich auf den Blatträndern außerhalb der Adern häufte. Die Ernährungsweise von A. fabae, Myzus persicae und M. ornatus wurde durch mikroskopische Suche nach Speichelscheiden in ganzen, aufgehellten Blättern sowie an Schnittserien untersucht. A. fabae saugt am Phloem der Blattadern, M. persicae jedoch zum Teil und M. ornatus fast völlig am Schwammparenchym.Es wird angenommen, daß dieses Gewebe neben dem Phloem eine weitere Nahrungsquelle darstellte welche von gewissen Blattläusen in unterschiedlichem Ausmaße ausgenutzt zu werden vermag. Unterschiede sowohl im Anteil der am Mesophyll saugenden Population wie in dem bevorzugten Gefäßdurchmesser der von den Blattläusen besogenen Adern sind ursächlich mit den artcharakteristischen Ansiedlungsweisen verbunden. Unterschiede in beiden Richtunger werden zwischen den Arten deutlich, und ähnliche Unterschiede treten zwischen den Alters-gruppen einiger Aphidenarten häufig auf. Dem Auftreten wechselweise nutzbarer Nahrungsquellen, die in etwas verschiedener Weise ausgebeutet werden können, wird potentielle Bedeutung im Hinblick auf die Virusübertragung sowie auf die Wirksamkeit systemischer Insektizide beigemessen.

     

A serological comparison of the three morphological phases of Coccidioides immitis; complement fixation tests with anti-Histoplasma capsulatum serum

Summary The results of this study indicated that antigens prepared from the three morphological phases ofCoccidioides immitis differed in their complement fixing activity with anti-Histoplasma capsulatum pooled serum. Spherule antigens were serologically less active in tests with the anti-H. capsulatum pooled serum than antigens prepared from arthrospores and from mycelium.Antigenic determinants which are common toC. immitis andH. capsulatum appeared to be located on the intact arthrospore cellular surface but not on the surface of spherule cells.Part of a dissertation submitted to the Graduate School of Duke University in partial fulfillment of requirements for the Ph. D. degree.This work was supported by contract with the Department of the Army, Fort Detrick, Frederick, Maryland.In conducting the research reported herein, the investigators adhered to Guide for Laboratory Animal Facilities and Care established by the Committee on the Guide for Laboratory Animal Facilities and Care of the Institute of Laboratory Animal Resources, NAS-NRC.