Biomedical subjects

I De Meester

Publications and source records attributed to I De Meester.

At least 55 records · Page 3Linked to original sources

PubMed1996

Distribution of prolyl oligopeptidase in human peripheral tissues and body fluids.

Prolyl oligopeptidase (EC 3.4.21.26) activity was measured in human tissue homogenates and body fluids. The enzyme was ubiquitously present, revealing high activity in renal cortex, epithelial cells, fibroblasts, testis, lymphocytes and thrombocytes. The activity in the body fluids was low. Prolyl oligopeptidase activity was significant higher in tumours of prostate, lung and sigmoid, than in the healthy tissues. Sera of individuals suffering from HIV infection, malaria, prostate cancer or benign prostate hypertrophy contained lowered activity. Interestingly, the low serum activity during prostate carcinoma increased upon medical treatment with anti-androgens. This suggests hormonal control of the gene transcript. A positive correlation with angiotensin converting enzyme activity in hypertensive patients was demonstrated and this further supports the possible involvement of prolyl oligopeptidase in the renin-angiotensin system and in the pathogenesis of hypertension.

Acquired Immunodeficiency Syndrome↗

PubMed1995

The purification, characterization and analysis of primary and secondary-structure of prolyl oligopeptidase from human lymphocytes. Evidence that the enzyme belongs to the alpha/beta hydrolase fold family.

Prolyl oligopeptidase was isolated and purified to homogeneity from human lymphocytes, yielding a specific activity of 7780 mU/mg. The molecular mass using size-exclusion chromatography matches the 76 kDa obtained by SDS/PAGE. This provides evidence that prolyl oligopeptidase is a monomer. The isoelectric point is 4.8 as judged by isoelectric focusing in free solution. Di-isopropyl fluorophosphate and phenylmethylsulphonyl fluoride completely abolish the activity, classifying the enzyme as a serine proteinase. The inhibition by p-chloromercuribenzoic acid indicates the importance of a free sulfhydryl group near the active-site. alpha 1-Casein and ornithine decarboxylase, two proteins containing a PEST sequence, inhibit prolyl oligopeptidase, but were not hydrolyzed. This demonstrates that prolyl oligopeptidase is not participating in the metabolism of proteins according to a PEST-dependent pathway. alpha 1-Antitrypsin partially inhibits the enzyme but in contrast, aprotinin does not. Its inability to cleave corticotropin-releasing factor, ubiquitin, albumin and aprotinin, together with the hydrolysis of bradykinin between Pro7-Arg8 confirms the affinity of prolyl oligopeptidase for small peptides. Multiple sequence alignment does not reveal any similarity with proteases of known tertiary structure. Secondary-structure prediction displays striking similarity with dipeptidyl peptidase IV and acylaminoacyl peptidase. Two characteristic features of the members of the prolyl oligopeptidase family of serine proteases are high-lighted: the linear arrangement of the catalytic triad is nucleophile-acid-base and the proteolytic cleavage releasing the catalytically active C-terminal region of around 500 amino acids from the N-terminal sequence. Secondary structure prediction and comparison of the active-site of serine proteinases with known three-dimensional coordinates prove that Asp641 is the third member of the catalytic triad. The secondary structural organization of the protease domain of prolyl oligopeptidase is in accordance with the alpha/beta hydrolase fold.

Amino Acid Sequence↗

PubMed1995

Proline motifs in peptides and their biological processing.

Many biologically important peptide sequences contain proline. It confers unique conformational constraints on the peptide chain in that the side-chain is cyclized back onto the backbone amide position. Inside an alpha-helix the possibility of making hydrogen bonds to the preceding turn is lost and a kink will be introduced. The conformational restrictions imposed by proline motifs in a peptide chain appear to imply important structural or biological functions as can be deduced from their often remarkably high degree of conservation as found in many proteins and peptides, especially cytokines, growth factors, G-protein-coupled receptors, V3 loops of the HIV envelope glycoprotein gp 120, and neuro- and vasoactive peptides. Only a limited number of peptidases are known to be able to hydrolyze proline adjacent bonds. Their activity is influenced by the isomeric state (cis-trans) as well as the position of proline in the peptide chain. The three proline specific metallo-peptidases (aminopeptidase P, carboxypeptidase P and prolidase) are activated by Mn2+, whereas the three serine type peptidases cleaving a post proline bond (prolyl oligopeptidase, dipeptidyl peptidase IV, and prolylcarboxypeptidase) share the sequential order of the catalytic Ser-Asp-His triade, which differentiates them from the chymotrypsin (His-Asp-Ser) and subtilisin (Asp-His-Ser) families. An endo or C terminal Pro-Pro bond and an endo pre-Pro peptide bond possess a high degree of resistance to any mammalian proteolytic enzyme.

Amino Acid Sequence↗

PubMed1994

Cloning and sequence analysis of the gene encoding human lymphocyte prolyl endopeptidase.

The human cDNA encoding prolyl endopeptidase, a cytoplasmic endoprotease which hydrolyses the peptide bond at the C-terminal side of proline, was sequenced. After the isolation of the 3' terminal fragment of the pep cDNA sequence from a human lymphocyte cDNA library, an approach based on the polymerase chain reaction (PCR) was undertaken to obtain the complete pep cDNA. Overlapping DNA fragments were generated by PCR from cDNA synthesized from human lymphocyte mRNA. The DNA fragments were subcloned and sequenced. The complete cDNA is 2562 nucleotides (nt) in length and contains an open reading frame coding for a protein of 710 amino acids (aa). Comparison of the primary PEP sequences from human lymphocyte and pig brain shows 97% identify. The aa sequence analysis shows homology with bacterial PEPs and with protease II from Escherichia coli. Asp641 probably participates in the active site of PEP.

Amino Acid Sequence↗

PubMed1994

Binding of adenosine deaminase to the lymphocyte surface via CD26.

The relationship between CD26/dipeptidyl peptidase IV, an ectopeptidase involved in T cell activation, and the binding protein for adenosine deaminase (ADAbp) was studied. Monoclonal antibodies (mAb) against CD26 and ADAbp, respectively, showed a similar binding profile on various lymphocyte subsets from the peripheral blood. The adenosine deaminase (ADA) itself blocked the binding of a specific set of anti-CD26 mAb (among these the anti-TA5.9 mAb) on lymphocytic CD26; ADA also hindered the binding of soluble CD26 to the same immobilized anti-CD26 mAb. In addition, the interaction between immobilized ADA and purified CD26/DPP IV was inhibited by the anti-TA5.9 mAb. ADA did not inhibit the specific peptidase activity of CD26. Neither soluble nor immobilized ADA was able to down-modulate CD26 on the lymphocyte surface. Our data thus confirm the identity between ADAbp and CD26 and identify some epitopes, crucial in the binding of ADA to CD26.

Adenosine Deaminase↗

PubMed1994

Purified and cell-bound CD26: enzymatic inhibition, antibody binding profile, and expression on T cells in relation to other surface markers.

The CD26 activation antigen (Ag), which is expressed on a subpopulation of human T cells, has been characterized as dipeptidyl peptidase IV (DPP IV, EC 3.4.14.5). We investigated some molecular and inhibition characteristics as well as the monoclonal antibody (mAb)-binding profile of this molecule purified from human lymphocytes. Among the antibodies we explored, two, anti-BT5/9 and anti-TA5.9, exhibited a high affinity for both purified and cell-bound CD26 Ag. Their significance in the study of immunologic memory and lymphocyte activation is discussed in relation to other markers of lymphocyte activation. Among 4 types of inhibitors studied, Pro-boroPro proved to be the most promising substance for further research on the physiological role of dipeptidyl peptidase IV (CD26).

Antibodies, Monoclonal↗

PubMed1994

Dipeptidyl peptidase IV: development, design, synthesis and biological evaluation of inhibitors.

Dipeptidyl peptidase IV (DPP IV, EC 3.4.14.5) is a highly specific serine protease which cleaves off N-terminal dipeptides from peptides with a penultimate proline or alanine. The enzyme is widely distributed in mammalian cells and tissues, but specific activities differ greatly. In the hematopoietic system it is found almost exclusively on T cells, where it is identified as CD26, a T cell activation molecule. DPP IV may be involved in the metabolism of peptides, intestinal assimilation and cell adhesion and it plays an integral role in T cell activation. DPP IV inhibitors may provide help during the further elucidation of the biological function(s) of this enzyme. Moreover, because of the integral role the enzyme plays in T cell activation, specific inhibition of DPP IV may constitute a new way of immune modulation. N-Peptidyl-O-acylhydroxylamines and boronic acid analogues of proline and alanine are two known DPP IV inhibitors. The major drawbacks for their therapeutic use are for the hydroxylamines, the toxicity and for the boronic acid derivatives, the chemical liability. A low toxicity, acceptable stability and a high specificity are essential criteria for the design of inhibitors that are suitable, not only for experimental, but also for therapeutic use. Therefore we proposed 5 types of potential DPP IV inhibitors: azapeptides, azetidines, Michael substrates, reduced peptides and phosphonic acids. All the synthesized compounds possess a substrate-like structure, which is a pre-requisite for recognition by the enzyme. We choose for a modified proline or alanine at the penultimate position, substituted with glycine, alanine, valine, isoleucine or phenylalanine at the N-terminus. The prepared compounds were screened biologically at a 5 mM concentration with a fluorometric method using Gly-Pro-4-Me-2NA as substrate.

Aza Compounds↗

PubMed1994

Components of biological variation in prolyl endopeptidase and dipeptidyl-peptidase IV activity in plasma of healthy subjects.

We investigated the components of biological variation in plasma prolyl endopeptidase (PEP; EC 3.4.21.26) and dipeptidyl-peptidase IV (DPP IV; EC 3.4..14.5) activity in healthy individuals. We took monthly blood samples from 26 healthy volunteers for determination of plasma PEP and DPP IV activity during one calendar year. The estimated CVs for PEP activity were: total (CVt) = 25.0%, interindividual (CVg) = 13.9%, and intraindividual (CVi) = 16.8%. There was a statistically significant (P < 0.0001) seasonal pattern in plasma PEP activity, with significantly higher values in the fall than in the other seasons. The peak-trough difference in the yearly variation in PEP activity, expressed as a percentage of the mean, was as high as 56.8%. The estimated CVs for DPP IV activity were: CVt = 17.1%, CVg = 14.5%, and CVi = 8.2%. DPP IV activity was significantly (P < 0.0001) higher in summer than in the other seasons but the amplitude of the yearly variation was small.

Adolescent↗

PubMed1993

Relationships between lower plasma L-tryptophan levels and immune-inflammatory variables in depression.

Despite much research, the pathophysiology underlying lower L-tryptophan (L-TRP) availability in major depression has remained elusive. The present study investigates whether lower L-TRP availability in major depression is related to immune activation which may occur in that illness and is known to modulate L-TRP metabolism. Toward this end, the authors have measured the following in depressed patients and normal control subjects: plasma levels of L-TRP, and the competing amino acids (CAA) valine, leucine, isoleucine, tyrosine, and phenylalanine, together with indices of immune function such as haptoglobin (Hp) and transferrin (Tf) plasma levels, dipeptidyl peptidase IV (DPP IV) serum activity, and mitogen-induced culture supernatant interleukin-6 (Il-6) production. Both plasma levels of L-TRP and the L-TRP/CAA ratio were significantly lower in major depressed subjects as compared with healthy control subjects. There were significant correlations between plasma L-TRP levels, on the one hand, and Tf plasma levels, DPP IV activity (both positive), Il-6 production, and Hp plasma levels (both negative), on the other. Up to 63.7% of the variance in L-TRP plasma concentrations could be explained by DPP IV, Hp, Il-6 values, and gender. Up to 50% of the variance in the L-TRP/CAA ratio could be explained by Hp values (negative correlation) and gender. It is hypothesized that lower plasma L-TRP availability in major depression may be related to the immune response in that illness.

Adult↗

PubMed1993

Decreased expression of the memory marker CD26 on both CD4+ and CD8+ T lymphocytes of HIV-infected subjects.

Using a novel anti-CD26 (or anti-dipeptidyl peptidase IV) monoclonal antibody, we showed that the absolute numbers and the proportions of T4 and T8 cells expressing CD26 were significantly lower in HIV-infected persons than in controls. The absolute number of CD26+ T4 cells decreased according to disease progression, whereas the number of CD26+ T8 cells was low throughout all clinical stages. These trends were similar in CD26 dim and bright positive T-cell subsets. In both controls and HIV-positive subjects, the CD26 bright positive T cells were restricted to the CD45RO+ subset and preferentially co-expressed CD25 but largely lacked HLA-DR and CD38. Recall antigen-responsive cells from seronegative individuals were shown to co-express CD26 and CD45RO. The deficient CD26 expression on T8 cells from HIV-infected subjects could be normally upregulated after in vitro stimulation. In contrast to decreased T-cell-bound CD26, the enzymatic activity of plasma CD26/dipeptidyl peptidase IV was unchanged in HIV-infected patients compared with controls. We conclude that HIV infection leads to a deficient in vivo co-expression of CD26 bright and CD45RO on T cells. We speculate that this deficiency might play a part in the decrease of immunological memory during HIV infection.

ADP-ribosyl Cyclase↗

PubMed1992

Characterization of dipeptidyl peptidase IV (CD26) from human lymphocytes.

The membrane-bound dipeptidyl peptidase IV (DPP IV, EC 3.4.14.5) has been purified 5,400-fold from human peripheral blood mononuclear cells. The purification procedure included detergent solubilization and successive chromatography on DEAE Sepharose Fast Flow, Con A Sepharose, Cu2+ loaded metal-chelating Sepharose, Sephacryl S-300 High Resolution and Q Sepharose Hiload. The molecular mass of the native, detergent solubilized enzyme estimated by gel filtration was 264.kDa. Chromatofocusing indicated a pI of approximately 5.0. The pI optimum was 8.7. The enzymatic activity of the purified preparation was irreversibly inhibited by N-(H-Phe-Pro)-O-(4-nitrobenzoyl)hydroxylamine hydrochloride in the micromolar range. The binding of purified DPP IV to CD26 monoclonal antibodies confirmed the identity between CD26 and dipeptidyl peptidase IV. The purification and characterization of lymphocytic dipeptidyl peptidase IV is of great value for the identification of its natural substrates and for the study of its physiological significance in the T-lymphocyte function.

Amino Acid Sequence↗

PubMed1992

Kininase activity in human platelets: cleavage of the Arg1-Pro2 bond of bradykinin by aminopeptidase P.

A proline-specific peptidase aminopeptidase P (APP, EC 3.4.11.9) that cleaves the Arg1-Pro2 bond of bradykinin was isolated from human platelets by liquid chromatography. The enzyme was purified 557 times. The native molecule has a M(r) of 223,000. Human platelet APP exists as a trimer with a subunit M(r) of 71,000. The apparent Km of platelet APP is 66 mumol/L for bradykinin and 47 mumol/L for the internally quenched fluorogenic substrate Lys (2,4-dinitrophenyl)-Pro-Pro-NH-CH2-CH2-NH-2-aminobenzoyl. 2HCl which is used for the routine determination of the enzyme activity. The optimum pH for hydrolysis of the fluorogenic substrate is 8.0, and the optimum temperature is 43 degrees. Platelet APP is inhibited by 1,10-phenanthroline and activated by Mn2+, thus confirming its metalloprotease nature. Cu2+, Zn2+ and Hg2+ are strongly inhibitory. Inhibition by cysteine protease inhibitors suggests the presence of a thiol group essential for enzymatic activity. Serine protease inhibitors do not affect the enzyme activity.

Aminopeptidases↗

PubMed1992

Disturbances in dexamethasone suppression test and lower availability of L-tryptophan and tyrosine in early puerperium and in women under contraceptive therapy.

This study investigates the function of the hypothalamic-pituitary-adrenal (HPA)-axis and the availability of L-tryptophan and tyrosine to the brain in postpartum women and in women taking long-term oral contraceptives. To this end, we have measured the following parameters in 50 women (i.e. 9 normal controls, 10 women taking oral contraceptives, and 31 postpartum females): plasma cortisol, L-tryptophan, tyrosine and the amino acids (CAA) known to compete with them for transport through the blood-brain barrier. We have determined the effects of 1 mg of dexamethasone on the above-mentioned biological markers in postpartum females. Plasma cortisol and tyrosine were significantly higher and lower, respectively, in puerperium and in women under contraceptive therapy as opposed to normal controls. L-Tryptophan was significantly lower in postpartum females, whilst the L-tryptophan/CAA ratio did not differ across the three study groups. Postpartum females revealed a significant negative relationship between the availability of L-tryptophan to the brain and postpartum mood, as measured by Zung's Depression and Anxiety Scales and State Anxiety Inventory. Dexamethasone had a significant suppressive effect on L-tryptophan/CAA and tyrosine/CAA ratios, with cortisol nonsuppression appearing in 82% of the postpartum females.

Adult↗

PubMed1992

Localization and characterization of aminopeptidase P in bovine adrenal medulla.

Aminopeptidase P (EC 3.4.11.9) is demonstrated for the first time in the cytosolic fraction of chromaffin cells of the bovine adrenal medulla. The enzyme is inhibited by metal chelators and by sulfhydryl-reactive agents, which suggests that both a tightly bound metal ion and a cysteine residue are necessary for enzymatic activity. Aminopeptidase P might be important for the modulation of the biological activity of neuropeptides. Its occurrence in the adrenal chromaffin cells provides a useful tool for studying the function of this unique proline-specific peptidase in neuropeptide processing and secretion.

Adrenal Medulla↗

PubMed1992

Distribution of proline-specific aminopeptidases in human tissues and body fluids.

The proline-specific peptidases, aminopeptidase P (EC 3.4.11.9) and dipeptidyl peptidase IV (EC 3.4.14.5), were measured in human tissue homogenates and physiological fluids. All tissues examined contained measurable aminopeptidase P and dipeptidyl peptidase IV activities. High specific activities for both enzymes under study were found in benign prostatic hypertrophy. Normal prostate and prostatic adenocarcinoma had a much lower activity. This difference, however, is not reflected in the serum values of the patients. The most striking finding is the extremely high activity of dipeptidyl peptidase IV in prostatosomes, prostate-derived organelles, which occur freely in human seminal plasma, and which are important for enhancement of sperm forward motility.

Adenocarcinoma↗

PubMed1992

A sensitive method for the assay of serum prolyl endopeptidase.

A new fluorimetric assay for the determination of prolyl endopeptidase (EC 3.4.21.26) was developed. The synthetic substrate Z-glycyl-prolyl-4-methylcoumarinyl-7-amide (0.2 mmol/l; 5 microliters), K-phosphate buffer (100 mmol/l, pH 7.5; 100 microliters), and serum (10 microliters) are incubated for 120 min at 37 degrees C. The reaction is stopped with acetic acid (1.5 mol/l; 500 microliters) and the released 7-amino-4-methylcoumarin is measured fluorimetrically. The mean value of prolyl endopeptidase catalytic activity concentration in serum for 120 healthy volunteers was 0.455 (SD = 0.092) mumol of 7-amino-4-methylcoumarin released per litre of serum per minute. The proposed procedure is sensitive, robust, and economical.

Adolescent↗

PubMed1991

Decreased serum dipeptidyl peptidase IV activity in major depression.

It has been recently shown that severe depression is characterized by immune dysfunctions such as blunted mitogen-induced blast transformation, which is linked to interleukin-2 (IL-2) mechanisms, and to autoimmune responses. In order to explore one of the putative pathophysiological mechanisms underlying both factors, we have measured the predexamethasone and postdexamethasone serum dipeptidyl-peptidase IV (DPP IV) activity in depressed inpatients and normal controls. This enzyme is an important mediator of IL-2-related blast proliferation, and it may play a role in autoimmunity. We found significantly lower DPP IV levels in major depressives as compared with healthy controls, and melancholics exhibited significantly lower enzyme activity than minor depressives. There was a significant negative correlation between serum DPP IV activity and the severity of illness. However, we were unable to detect any significant relationships between DPP IV on the one hand, and mitogen-induced blast transformation, soluble IL-2 receptor accumulation in PHA culture supernatant, total number of leukocytes and lymphocytes, T lymphocytes, CD4+ and CD25+ cells, on the other. Men exhibited significantly higher serum DPP IV levels than women.

Adjustment Disorders↗

PubMed1991

Aminopeptidase P and dipeptidyl peptidase IV activity in human leukocytes and in stimulated lymphocytes.

Human white blood cells were shown to contain high aminopeptidase P activity. The specific activities found in the high-speed supernatant of the extracts of granulocytes, lymphocytes and monocytes ranged from 30 to 70 units per mg protein. Culturing lymphocytes during 7 days in the presence of phytohaemagglutinin resulted in a 70-200% increase in the specific aminopeptidase P activity and a 200% increase in the specific activity of dipeptidyl peptidase IV. The time-course of the activity of both aminopeptidase P and dipeptidyl peptidase IV during the stimulation of human T-lymphocytes by phytohaemagglutinin indicates an involvement of these two enzymes in the proliferative process of these immunocompetent cells. Due to their substrate specificity their potential substrates must have the N-terminal Xaa-Pro sequence known to be present in several immunologically important polypeptides.

Aminopeptidases↗