Assignment of the human gamma-glutamyl transferase gene to the long arm of chromosome 22

Summary We have determined the chromosomal location of the human gene for gamma-glutamyltransferase (GGT). This study was done by in situ hybridization of human metaphase spreads with a rat cDNA probe specific for this enzyme and constructed from two clones previously characterized in our laboratory. The final construct had a 1.6-kb-long insert covering 92% of the coding sequence for GGT. The new insert was also freed of any GC tails introduced for the cDNA cloning, because we observed that these sequences were responsible for a high background. Using this probe for the analysis of 136 human metaphase spreads, we observed a strong specific signal on chromosome 22 at the interface of q111-112 and a minor peak in q131. Thus GGT might represent a new marker for the study of certain diseases which have chromosomal abnormalities at these loci.     

gamma-Glutamyl transpeptidase: a single copy gene in the rat and a multigene family in the human genome

gamma-Glutamyl transpeptidase (GGT) genomic sequences were isolated from rat and human libraries using a rat GGT cDNA as a cross-species hybridization probe. Characterization of the human GGT clones by restriction mapping clearly establishes that at least four different GGT genes or pseudogenes are present in the human genome. All the rat genomic clones cover a 12.5-kilobase sequence and exhibit a unique restriction pattern. A precise quantitation of the rat GGT gene copy number by Southern blot analysis demonstrates that this sequence is present as a single copy/rat haploid genome. Therefore, the GGT gene organization is different between rat and human species; this raises the possibility of different regulatory mechanisms in the two species.     

Phosphorylation of β-lactoglobulin using amino acids as the sole base and nucleophile of the reaction

β-Lactoglobulin was phosphorylated with 20, 40, and 80 mol of POCl₃/mol protein in the presence of 4, 5, and 6 molar excess of basic amino acid per mol POCl₃. Maximal phosphorylation yields of 5 and 3 mol P/mol protein were achieved when the highest stoichiometries of POCl₃/arginine and lysine were used. Proportional high amounts of basic amino acids were also grafted to the protein molecule during its phosphorylation through the phosphoamide bond. Modified proteins displayed increased negative charges and reduced isoelectric points and were monomeric. The phosphorylated and phosphoamidatedβ-lactoglobulin showed improved functional properties.     

Solubility and reactivity of caseins and β-lactoglobulin in protic solvents

The study of the solubility of unstructured proteins (α_S1-, β-, and κ-casein) and well-structured globulin (β-lactoglobulin) in low water binary solvent systems demonstrated the crucial importance of solvent polarity and neutralization of protein polar functions on the final outcome of solubility experiments. The solubilities up to 38, 56, and 96% in CHCl₃/CH₃OH (1/1, v/v) acidified with HCl and up to 5, 10, and 25% in CHCl₃/CH₃OH (1/1, v/v) in the presence of triethylamine (TEA) were obtained for κ-, α_S1-, and β-casein, respectively. The importance of protein charge neutralization was apparent when the solubilization was performed in basified CHCl₃/CH₃OH media, giving the optimal results when the studied proteins were brought before to their isoionic point. The maximum solubility of β-casein at its pI in 30–70% methanol in CHCl₃ was reaching 50–60% with triethylamine (TEA) added. β-lactoglobulin could be solubilized up to 70% in CHCl₃/CH₃OH (7/3, v/v) acidified with HCl and up to 40% in CHCl₃/CH₃OH (3/7, v/v) in the presence of TEA. The observed yield of reductive alkylation of β-lactoglobulin was much higher (98%) when performed in studied solvent system than in aqueous conditions (75%). Apparently, steric hindrance of the well-folded β-barrel (in aqueous conditions) structure masks the portion of ε-NH₂ groups. In the case of unstructured aqueous media β-casein, 90% alkylation yields were obained in organic and aqueous conditions.     

Glucocorticoid hormones increase the activity of γ-glutamyltransferase in a highly differentiated hepatoma cell line

γ-Glutamyltransferase activity was detected in the plasma membrane of the highly differentiated hepatoma cell line Fao, (0.93 mU/mg cell protein). Dexamethasone (1 μM) provoked a 2–3-fold increase in the activity of the enzyme in the presence of fetal calf serum. Maximal induction occurred 48–72 h after addition of the glucocorticoid to the cell culture medium. The hormonal specificity was demonstrated by the relative potencies of several glucocorticoids and sex steroids: hydrocortisone and corticosterone increased γ-glutamyltransferase activity while tetrahydrocorticosterone and all sex steroids tested were ineffective. The effect of dexamethasone on γ-glutamyltransferase activity was specific since the activities of several other plasma membrane enzymes were not modified. The mechanism of the dexamethasone-induced increase in γ-glutamyltransferase activity was neither by modification of the affinity of the enzyme for its substrates nor by alteration of the subcellular distribution of the enzyme. This increase was prevented by cycloheximide and actinomycin D. The data presented are consistent with a specific glucocorticoid receptor-mediated induction of γ-glutamyltransferase activity in Fao cells. The kinetic parameters of the induction process by glucocorticoids are very similar to those found in adult rat liver. These results suggest that the Fao cell line is a very convenient system for the study of the molecular mechanisms of glucocorticoid effects on differentiated cells.     

The status and conservation of the Cape Gannet Morus capensis

The Cape Gannet Morus capensis is one of several seabird species endemic to the Benguela upwelling ecosystem (BUS) but whose population has recently decreased, leading to an unfavourable IUCN Red List assessment. Application of ‘JARA’ (‘Just Another Red-List Assessment,’ a Bayesian state-space tool used for IUCN Red List assessments) to updated information on the areas occupied by Cape Gannets and the nest densities of breeding birds at their six colonies, suggested that the species should be classified as Vulnerable. However, the rate of decrease of Cape Gannets in their most-recent generation exceeded that of the previous generation, primarily as a result of large decreases at Bird Island, Lambert’s Bay, and Malgas Island, off South Africa’s west coast (the western part of their range). Since the 1960s, there has been an ongoing redistribution of the species from northwest to southeast around southern Africa, and ～70% of the population now occurs on the south coast of South Africa, at Bird Island in Algoa Bay, on the eastern border of the BUS. Recruitment rather than adult survival may be limiting the present population; however, information on the seabird’s demographic parameters and mortality in fisheries is lacking for colonies in the northern part of the BUS. Presently, major threats to Cape Gannet include: substantially decreased availability of their preferred prey in the west; heavy mortalities of eggs, chicks and fledglings at and around colonies, inflicted by Cape Fur Seals Arctocephalus pusillus and other seabirds; substantial disturbance at colonies caused by Cape Fur Seals attacking adult gannets ashore; oiling; and disease.