Leucine aminopeptidase as an echo-enzyme of polymorphonuclear neutrophils

Intact polymorphonuclear neutrophils were modified chemically by a poorly permeable reagent, diazotized sulfanilic acid, and the changes in the activity of 5'-nucleotidase, alkaline phosphodiesterase, and leucine aminopeptidase were examined. Among three plasma membrane enzymes, 5'-nucleotidase activity was hardly detected in the human neutrophils. The activity of alkaline phosphodiesterase was observed in all the neutrophils examined, but was not inhibited by diazotized sulfanilic acid in the guinea-pig neutrophils. On the other hand, the activity of leucine aminopeptidase was not only found but also inhibited by diazotized sulfanilic acid without the inhibition of lactate dehydrogenase, a cytosol enzyme, in all the neutrophils, suggesting that leucine aminopeptidase is located generally on the plasma membrane as an ecto-enzyme in the neutrophils.     

Leucine aminopeptidase as an ecto-enzyme of polymorphonuclear neutrophils

Intact polymorphonuclear neutrophils were modified chemically by a poorly permeable reagent, diazotized sulfanilic acid, and the changes in the activity of 5′-nucleotidase, alkaline phosphodiesterase, and leucine aminopeptidase were examined. Among three plasma membrane enzymes, 5′-nucleotidase activity was hardly detected in the human neutrophils. The activity of alkaline phosphodiesterase was observed in all the neutrophils examined, but was not inhibited by diazotized sulfanilic acid in the guinea-pig neutrophils. On the other hand, the activity of leucine aminopeptidase was not only found but also inhibited by diazotized sulfanilic acid without the inhibition of lactate dehydrogenase, a cytosol enzyme, in all the neutrophils, suggesting that leucine aminopeptidase is located generally on the plasma membrane as an ecto-enzyme in the neutrophils.     

Leucine aminopeptidase in extracts of swine muscle

1. Leucine aminopeptidase (EC 3.4.1.1) has been demonstrated in swine muscle at a level of activity one-fifth that of the swine kidney. 2. The enzyme has been purified 110-fold by precipitation with ammonium sulphate, heat treatment and chromatography on Sephadex G-100. 3. The enzyme is heat-stable, but is rapidly inactivated below pH7. It requires Mg(2+) or Mn(2+) for activity. The Michaelis constant for leucine amide with Mg(2+)-activated enzyme is 5.0x10(-3)m. 4. Muscle leucine aminopeptidase is very similar to the kidney enzyme.     

Intracellular enzymatic response of lymphocytes and neutrophils in patients with cancer of the larynx.

In 20 men, aged 35 to 55 years, with untreated cancer of the larynx activity of lysosomal acid phosphatase (AP), beta-glucuronidase (GR) and N-acetyl-beta-glucosaminidase was determined cytochemically in peripheral blood lymphocytes and neutrophils by means of Barka and Anderson, Hayashi et al. and Hayashi's method, respectively; the results obtained were compared with those in 20 healthy men aged 20 to 30 years. Total count of GR-positive lymphocytes was higher in the patients than in normal persons. Total counts of AP-, GR-, and GS-positive lymphocytes with not disrupted enzyme-positive lysosomal granules within the cell cytoplasm were significantly lower and total counts of cells exhibiting the disruption of lysosomal granules and the diffuse type of cytochemical reaction were significantly higher in the patients when compared with the control group. The response of neutrophils consisted of a significant elevation in numbers of AP-, and GS-positive cells; overall score of enzyme activity studied in neutrophils was not altered in the patients. The authors disucss the significance of their observations in the light of data on participation of lymphocytic and neutrophilic lysosomal apparatus in the immunological response against tumour specific antigen in patients with cancer.     

Leucine aminopeptidase activity in pain-induced emotional stress

It is established that leucine aminopeptidase activity during emotional-algesic stress increases in the brain hemispheres, left ventricle and liver of the rats as compared to that of intact animals. Maximum activation of the enzyme in the brain and liver is detected for the first two days after the stress while in the heart - for the whole period of the total stress damages volume development. A conclusion is drawn that the shifts observed in leucine aminopeptidase activity during emotional-algesic stress affect the methionine-enkephalin and leucin-enkephalin ratio.     

Leucine aminopeptidase and hatching of Schistosoma mansoni eggs

Leucine aminopeptidase (LAP) activity has been measured in extracts of eggs, miracidia, cercariae and adult worms of Schistosoma mansoni. Activity measured at pH 7.2 using L-leu-7-amino-4-trifluoro-methylcoumarin as substrate is 6- to 17-fold greater in eggs than in other life stages. LAP activity is also high in soluble egg antigen preparations and in hatching fluid. The release of LAP from eggs parallels hatching, and inhibitors of LAP also inhibit hatching. The possible role of LAP in the hatching process of S. mansoni eggs is discussed.     

Leucine aminopeptidase and ferricyanide reductase activities in radish microsomes

A NADH-ferricyanide reductase activity found in radish microsomes isolated from germinated seeds has been shown to be stimulated by pCMB and pCMBS which are both strong nactivators of many plant proteolytic enzymes. In the same preparation a leucine aminopeptidase was found while endoprotease and carboxypeptidase activities were not detected using exogenous substrates. The aminopeptidase, highly active at the same optimal pH-condition of FeCN reductase, was stimulated by CoCl₂ and non-polar detergents (Triton X-100 and Brij 35). It was inhibited by sulphydryl reagents. By gel filtration of microsomal detergent extract two peaks of activity were separated: red I coeluted with LeuAPase and red II, free of aminopeptidase. Red I, a protein, was inhibited by sulphydral reagents and stimulated by duroquinone. Red II, stimulated by pCMB, is not a protein because of the small size and the noninfluence of heating treatment on catalytic activity.     

Leucine aminopeptidase in exsheathing fluid of North American and Australian Haemonchus contortus

Rogers W. P. and Brooks F. 1978. Leucine aminopeptidase in exsheathing fluid of north American and Australian Haemonchus contortus. International Journal for Parasitology8: 55–58. Juveniles of Haemonchus contortus from north America and Australia produced exsheathing fluid containing leucine aminopeptidase when stimulated in tetraborate-carbon dioxide medium. Exsheathment in this medium was inhibited by 1, 10-phenanthroline, 10^?3M, and this inhibition was largely reversed by Zn²⁺, 10^?3M. This supports the view that the enzyme is produced by the juveniles and that it is concerned in exsheathment.     

Leucine aminopeptidase and exsheathing activity in preparations from Haemonchus contortus

Rogers W.P. and Brooks F. 1978. Leucine aminopeptidase and exsheathing activity in preparations from Haemonchus contortus. International Journal for Parasitology 8: 449–452. Exsheathing activity relative to leucine aminopeptidase activity (LAP) was greater in exsheathing fluid of infective juveniles of Haemonchus contortus than extracts of homogenates of the same organism. In both preparations the biological and enzyme activities were precipitated with acetone 20 v/v and ammonium sulphate, 40% saturation. Broad peaks of exsheathing and LAP activities obtained by sucrose density-gradient centrifugation and on Sephadex G150 overlapped but the peak of biological activity was always found on the low mol. wt. side of the LAP peak. LAP in exsheathing fluid was separated into two sharp peaks in polyacrylamide gradient-pore electrophoresis. In four experiments the major peak gave a mol. wt. within the limits 345,000–354,500. A minor peak was obtained at 1,800,000. Exsheathing activity remained broadly distributed but fell mostly on the low mol. wt. side of the major LAP peak.It is concluded that LAP cannot be the sole agent involved in exsheathment a lipase may be necessary to expose the substrate attacked by LAP.     

Effect of proper-myl on humoral interactions between activated lymphocytes and neutrophils in patients with myeloproliferative diseases

The lymphokine synthesizing function of peripheral blood lymphocytes (PBL) was studied in 22 patients with chronic myeloid leukemia (CML), 28 patients with subleukemic myelosis (SLM) and 15 healthy persons. The index of the neutrophil stimulation (INS) in CML (1.73 +/- 0.068) and SLM (1.458 +/- 0.004) was statistically significantly lower than that in the healthy persons (2.6 +/- 0.07). The use of proper-myl in the complex therapy markedly increased the ability of PBL to produce the factor stimulating phagocytic activity of neutrophils (INS 1.94 +/- 0.04 and 1.832 +/- 0.092). On the basis of the findings it was recommended to use proper-myl for prevention and treatment of infectious complications.     

Leucine aminopeptidase of the human blood flukes, Schistosoma mansoni and Schistosoma japonicum

An array of schistosome endoproteases involved in the digestion of host hemoglobin to absorbable peptides has been described, but the exoprotease responsible for catabolising these peptides to amino acids has yet to be identified. By searching the public databases we found that Schistosoma mansoni and Schistosoma japonicum express a gene encoding a member of the M17 family of leucine aminopeptidases (LAPs). A functional recombinant S. mansoni LAP produced in insect cells shared biochemical properties, including pH optimum for activity, substrate specificity and reliance on metal cations for activity, with the major aminopeptidase activity in soluble extracts of adult worms. The pH range in which the enzyme functions and the lack of a signal peptide indicate that the enzyme functions intracellularly. Immunolocalisation studies showed that the S. mansoni LAP is synthesised in the gastrodermal cells surrounding the gut lumen. Accordingly, we propose that peptides generated in the lumen of the schistosome gut are absorbed into the gastrodermal cells and are cleaved by LAP to free amino acids before being distributed to the internal tissues of the parasite. Since LAP was also localised to the surface tegument it may play an additional role in surface membrane re-modelling.     

Leucine aminopeptidase (LAP) activity and sexual dimorphism in rat exorbital lacrimal gland

Leucine amonopeptidase (LAP) activity was histochemically studied in the rat exorbital lacrimal gland. The enzyme is present in the secreting cells of both male and female prepuberal rats, in vivo and in monolayer cultures, while in the adult rat it is demonstrable only in the female. Furthermore, LAP is influenced by the sex hormones, disappearing in the female gland after testosterone treatment and appearing in the adult male gland after administration of estradiol or cyproterone acetate. These results show that the behaviour of LAP can be considered as another character of sexual dimorphism of the rat exorbital gland. Furthermore, our findings, showing an inverse relationship between LAP activity and the PAS positivity of the secretion products, suggest the hypothesis that the presence of this exopeptidase could induce qualitative modifications of one or more secreted glycoprotein.