scholarly journals Purification and properties of a human seminal proteinase

1972 ◽  
Vol 126 (5) ◽  
pp. 1135-1140 ◽  
Author(s):  
Frank N. Syner ◽  
Kamran S. Moghissi
Keyword(s):  
Ethyl Ester ◽  
Seminal Plasma ◽  
Gel Filtration ◽  
Enzyme Assays ◽  
Human Seminal Plasma ◽  
Ph Optimum ◽  
Purification And Properties ◽  
Ammonium Sulphate Fractionation ◽  
Substrate Concentrations

1. A method is described for the purification of a proteinase, present in human seminal plasma and previously shown to accelerate migration of spermatozoa through cervical mucus in vitro. A 25-fold purification was achieved in three steps, consisting of ammonium sulphate fractionation, chromatography on CM-cellulose and gel filtration. 2. The enzyme displays some properties similar to chymotrypsin: pH optimum 7.5–8.0; substrate preference of casein, haemoglobin and benzoyltyrosine ethyl ester but not benzoylarginine ethyl ester; mol.wt. 33000. However, it is unaffected by 1mm-di-isopropyl phosphofluoridate or 1mm metal cations, and in this respect differs from chymotrypsin. 3. The properties of the enzyme strongly resemble those of the ‘chymotrypsin-like’ enzyme discovered in seminal plasma by Lundquist et al. (1955). 4. The use of dimethyl-casein permitted the performance of enzyme assays at substrate concentrations five times higher (up to 50mg/ml) than could be achieved with ordinary casein (10mg/ml).

Partial purification and properties of an L-arabinose dehydrogenase from Azospirillum brasiliense

1983 ◽  
Vol 29 (2) ◽  
pp. 242-246 ◽  
Author(s):  
Norman J. Novick ◽  
Max E. Tyler
Keyword(s):  
Molecular Weight ◽  
Divalent Cation ◽  
Gel Filtration ◽  
Reducing Agent ◽  
Partial Purification ◽  
Ph Optimum ◽  
Glycine Buffer ◽  
Purification And Properties ◽  
Filtration Technique

An L-arabino-aldose dehydrogenase responsible for the oxidation of L-arabinose to L-arabino-γ-lactone has been purified 59-fold from L-arabinose grown cells of Azospirillum brasiliense. The dehydrogenase was found to be specific for substrates with the L-arabino-configuration at carbons 2, 3, and 4. Km values for L-arabinose of 75 and 140 μM were found with NADP and NAD as coenzymes, respectively. The enzyme had a pH optimum of 9.5 in glycine buffer and was stable when heated to 55 °C for 5 min. No enhancement of activity in the presence of any divalent cation or reducing agent tested was found. L-Arabinose dehydrogenase had a molecular weight of 175 000 as measured by the gel filtration technique.

scholarly journals Purification and properties of l-mandelate dehydrogenase and comparison with other membrane-bound dehydrogenases from Acinetobacter calcoaceticus

1987 ◽  
Vol 248 (3) ◽  
pp. 871-876 ◽  
Author(s):  
M E Hoey ◽  
N Allison ◽  
A J Scott ◽  
C A Fewson
Keyword(s):  
Lactate Dehydrogenase ◽  
Ion Exchange ◽  
Gel Filtration ◽  
Acinetobacter Calcoaceticus ◽  
Ion Exchange Chromatography ◽  
Exchange Chromatography ◽  
Ph Optimum ◽  
Purification And Properties ◽  
Potential Inhibitors ◽  
Triton X

L-Mandelate dehydrogenase was purified from Acinetobacter calcoaceticus by Triton X-100 extraction from a ‘wall + membrane’ fraction, ion-exchange chromatography on DEAE-Sephacel, (NH4)2SO4 fractionation and gel filtration followed by further ion-exchange chromatography. The purified enzyme was partially characterized with respect to its subunit Mr (44,000), pH optimum (7.5), pI value (4.2), substrate specificity and susceptibility to various potential inhibitors including thiol-blocking reagents. FMN was identified as the non-covalently bound cofactor. The properties of L-mandelate dehydrogenase are compared with those of D-mandelate dehydrogenase, D-lactate dehydrogenase and L-lactate dehydrogenase from A. calcoaceticus.

scholarly journals 229.Seminal plasma TGFβ activates pro-inflammatory cytokine synthesis in human cervical epithelial cells

2004 ◽  
Vol 16 (9) ◽  
pp. 229 ◽  
Author(s):  
D. J. Sharkey ◽  
S. A. Robertson
Keyword(s):  
Epithelial Cells ◽  
Seminal Plasma ◽  
Model Systems ◽  
Human Seminal Plasma ◽  
Neutralising Antibodies ◽  
Active Constituents ◽  
Gm Csf ◽  
Cytokine Synthesis ◽  
Cervical Epithelial Cells

Exposure to semen at intercourse in women elicits an inflammation-like response characterised by recruitment of inflammatory cells and expression of pro-inflammatory cytokines including GM-CSF, interleukin-6 (IL-6) and IL-8 (1). Studies in animal models have implicated TGFβ as the major active moiety in seminal plasma, and we have shown previously that TGFβ1 and TGFβ3 are present in high concentrations in human seminal plasma (>100 ng/mL), while TGFβ2 is less abundant. To investigate the physiological significance of each of the three TGFβ isoforms as pro-inflammatory agents in human seminal plasma, we have established in vitro model systems to measure human cervical cell cytokine synthesis. Primary cervical epithelial cells prepared from ectocervix of hysterectomy tissues or transformed Ect1 cells were incubated for 12 h with human recombinant TGFβ (isoforms 1, 2 or 3) or with seminal plasma in the presence or absence of isoform-specific TGFβ neutralising antibodies. Epithelial cell supernatants were recovered 24 h later and supernatants were analysed by commercial ELISA to quantify GM-CSF, IL-6 and IL-8 production. Each of the three TGFβ isoforms mimicked seminal plasma and were comparable in their capacity to stimulate >10-fold increases in both GM-CSF and IL-6 expression in a dose-responsive manner. In contrast, unlike seminal plasma none of the TGFβ isoforms induced IL-8 expression. Addition of neutralising antibodies to TFGβ1, TGFβ2 and TGFβ3 each effected >50% reduction in the ability of seminal plasma to induce GM-CSF and IL-6, but did not impair seminal plasma-stimulated IL-8 production. Together these data show that TGFβ1, TGFβ2 and TGFβ3 are major active constituents of seminal plasma, acting to elicit GM-CSF and IL-6 production in cervical epithelial cells. However, TGFβ does not fully account for the pro-inflammatory effects of human seminal plasma, and other active constituents remain to be identified. (1) D. J. Sharkey et al. (2003) Proc. SRB.

scholarly journals Purification and properties of acid phosphatase from Avena elatior L. seeds

2015 ◽  
Vol 46 (3) ◽  
pp. 481-488 ◽  
Author(s):  
E. Wieczorek ◽  
I. Lorenc-Kubis ◽  
B. Morawiecka
Keyword(s):  
Acid Phosphatase ◽  
Phosphatase Activity ◽  
Column Chromatography ◽  
Gel Filtration ◽  
Ph Optimum ◽  
Purification And Properties ◽  
Sepharose 4B

Acid phosphatase F1 from <i>Avena elatior</i> seeds was isolated and partially purified by means of alcohol precepitation, DEAE-, CM-column chromatography, Sephadex G-150, Sephadex G-200 and Sepharose 4B - gel filtration. The enzyme was stable at 50°C, pH 5.1. The pH optimum for phosphatase activity was 4.2. Fluoride, Zn<sup>2+</sup>, molybdate were effective inhibitors. EDTA and l, 10-phenanthroline activated the enzyme.

Purification and Properties of the Soluble 17β-Hydroxysteroid Dehydrogenase of Rabbit Uterus

1979 ◽  
Vol 34 (9-10) ◽  
pp. 726-737 ◽  
Author(s):  
Kunhard Pollow ◽  
Walter Eiger ◽  
Herrmann Heßlinger ◽  
Barbara Pollow
Keyword(s):  
Enzyme Activity ◽  
Dehydrogenase Activity ◽  
Gel Filtration ◽  
Deae Cellulose ◽  
Hydroxysteroid Dehydrogenase ◽  
Maximal Velocity ◽  
Ammonium Sulfate Precipitation ◽  
Ph Optimum ◽  
Mobility Data ◽  
Purification And Properties

Abstract 17 β-Hydroxysteroid dehydrogenase activity towards estradiol-17 β has been demonstrated in the 105,000 X g supernatant of rabbit uterus. Hydroxylapatite chromatography of the enzyme activity isolated by ammonium sulfate precipitation, gel filtration and DEAE-cellulose chromato­graphy yielded a single 17 β-hydroxysteroid dehydrogenase activity. Further purification of the enzyme preparation by isoelectric focusing resulted in multiple peaks of activity. The molecular weight or the enzyme, calculated from mobility data on Sephadex gel, is approximately 64,000. Some properties of partially purified 17 β-hydroxysteroid dehydrogenase activity have been studied. Estradiol-17 β reacts at a faster rate than testosterone. The Km for estradiol is 4.16X 10-5 mol/1 for the NAD-linked enzyme activity and 4.37 X 10-5 mol/1 when NADP as cofactor was used. The ratio of the maximal velocity for NADP to that for NAD was 1.42. The pH-optimum for estradiol appears between 9.5 and 10.5 and for estrone between 5.5 and 6.5. The enzyme appears to be of the sulfhydryl type.

In vitro studies of human seminal plasma allergy

1980 ◽  
Vol 66 (2) ◽  
pp. 148-154 ◽  
Author(s):  
J KOOISTRA ◽  
J YUNGINGER ◽  
P SANTRACH ◽  
J CLARK
Keyword(s):  
Seminal Plasma ◽  
In Vitro Studies ◽  
Human Seminal Plasma
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