Gene/Protein Disease Symptom Drug Enzyme Compound
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Query: EC:4.2.2.7 (heparinase)
 1,270 document(s) hit in 31,850,051 MEDLINE articles (0.00 seconds)

Pentosanpolysulfate (PPS) represents the product obtained after sulfation of xylan and is composed of beta 1----4-D-xylopyranose residues sulfated at C2 and C3. Studies have shown that this compound can often be effective in relieving the symptoms of interstitial cystitis (IC). In order to elucidate the mode of action of PPS in IC, a sensitive and reliable assay was needed. To this end we prepared an immunogenic form of PPS by coupling it to methylated bovine serum albumin (MBSA). This complex was used to immunize NZW rabbits (1 mg, IM). Four of five animals responded with anti-PPS antibodies, three of which had high titer (greater than 1/2000) as measured by an enzyme-linked immunosorbent assay (ELISA). All sera were routinely absorbed with an MBSA-Sepharose immunoadsorbent to remove anti-MBSA antibodies. ELISA inhibition tests were used to determine the sensitivity and specificity of the sera. At least 50 ng/ml of PPS could be routinely detected by this assay. A number of naturally occurring proteoglycans, polysaccharides, monosaccharides and disaccharides were examined for reactivity with the antibodies but only heparin was an effective inhibitor. Absorption with heparin immunoadsorbents reduced, but did not eliminate, the ability of heparin to inhibit anti-PPS binding. This activity could be destroyed by treatment with heparinase without affecting PPS inhibition. Normal urine did not affect the ELISA or ELISA inhibition tests and thus allowed the determination of PPS levels in IC patient urines. Initial analysis of seven IC patients receiving oral PPS revealed urine concentration of 0.8-16.0 micrograms/ml. No inhibition could be detected in pre-treatment urine samples.
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PMID:The production of antibodies to pentosanpolysulfate (ELMIRON, SP-54). 170 2

Whereas the glycocalyx of endothelial cells has been shown to influence solute flux from capillary microvessels, little is known about its contribution to the movement of macromolecules across the walls of other microvessels. We evaluated the hypothesis that a glycocalyx contributes resistance to protein flux measured in coronary arterioles. Apparent solute permeability (P(s)) to two proteins of different size and similar charge, alpha-lactalbumin (alpha-lactalb) and porcine serum albumin (PSA), was determined in arterioles isolated from the hearts of 43 female Yucatan miniature swine. P(s) was assessed in arterioles with an "intact" glycocalyx under control conditions and again after suffusion with adenosine (Ado, 10(-5) M, n = 42 arterioles, N = 29 pigs). In a second set of experiments (n = 21 arterioles, N = 21 pigs) arteriolar P(s) was determined before and after perfusion with enzyme (pronase or heparinase), which was used to digest the glycocalyx. P(s) was assessed a third time on those microvessels after exposure to Ado. Consistent with the hypothesis, P(s) for PSA (P(PSA)(s)) and P(s) for alpha-lactalb (P(alpha-lactalb)(s)) increased from basal levels following enzyme treatment. Subsequent suffusion with Ado, a significant metabolite known to alter coronary vascular smooth muscle tone and permeability, resulted in a significant reduction of basal P(alpha-lactalb)(s) in both untreated and enzyme-treated arterioles. Furthermore, in untreated arterioles, P(PSA)(s) was unchanged by Ado suffusion, whereas Ado induced a pronounced reduction in P(PSA)(s) of enzyme-treated vessels. These data demonstrate that in intact coronary arterioles an enzyme-sensitive layer, most likely at the endothelial cell surface, contributes significantly to net barrier resistance to solute flux.
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PMID:Role of a glycocalyx on coronary arteriole permeability to proteins: evidence from enzyme treatments. 1074 12

Using competitive binding experiments, it was found that native type XI collagen binds heparin, heparan sulfate, and dermatan sulfate. However, interactions were not evident with hyaluronic acid, keratan sulfate, or chondroitin sulfate chains over the concentration range studied. Chondrocyte-matrix interactions were investigated using cell attachment to solid phase type XI collagen. Pretreatment of chondrocytes with either heparin or heparinase significantly reduced attachment to type XI collagen. Incubation of denatured and cyanogen bromide-cleaved type XI collagen with radiolabeled heparin identified sites of interaction on the alpha1(XI) and alpha2(XI) chains. NH(2)-terminal sequence data confirmed that the predominant heparin-binding peptide contained the sequence GKPGPRGQRGPTGPRGSRGAR from the alpha1(XI) chain. Using rotary shadowing electron microscopy of native type XI collagen molecules and heparin-bovine serum albumin conjugate, an additional binding site was identified at one end of the triple helical region of the collagen molecule. This coincides with consensus heparin binding motifs present at the amino-terminal ends of both the alpha1(XI) and the alpha2(XI) chains. The contribution of glycosaminoglycan-type XI collagen interactions to cartilage matrix stabilization is discussed.
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PMID:Characterization of type XI collagen-glycosaminoglycan interactions. 1108 37

An approach is presented for the stable covalent immobilization of proteins with a high retention of biological activity. First, chemical modification studies were used to establish enzyme structural and functional properties relevant to the covalent immobilization of an enzyme to agarose based supports. Heparinase was used as a model enzyme in this set of studies. Amine modifications result in 75-100% activity loss, but the effect is moderated by a reduction in the degree of derivatization. N-hydroxysuccinimide, 1,1,1-trifluoroethanesulfonic acid, and epoxide activated agarose were utilized to determine the effect of amine reactive supports on immobilized enzyme activity retention. Cysteine modifications resulted in 25-50% loss in activity, but free cysteines were inaccessible to either immobilized bromoacetyl or p-chloromercuribenzoyl groups. Amine reactive coupling chemistries were therefore utilized for the covalent immobilization of heparinase. Second, to ensure maximal stability of the immobile protein-support linkage, the identification and subsequent elimination of the principal sources of protein detachment were systematically investigated. By using high-performance liquid chromatography (HPLC), electrophoresis, and radiolabeling techniques, the relative contributions of four potential detachment mechanisms-support degradation, proteolytic degradation, desorption of noncovalently bound protein, and bond solvolysis-were quantified. The mechanisms of lysozyme, bovine serum albumin, and heparinase leakage from N-hydroxysuccinimide or 1,1,1-trifluoroethanesulfonic acid activated agarose were elucidated. By use of stringent postimmobilization support wash procedures, noncovalently bound protein loss. An effective postimmobilization washing procedure is presented for the removal of adsorbed protein and the complete elimination of immobilized protein loss.
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PMID:An approach for the stable immobilization of proteins. 1859 60