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Libia Sanz

Publications and source records attributed to Libia Sanz.

25 records · Page 2Linked to original sources
PubMed2004

Snake venomics: characterization of protein families in Sistrurus barbouri venom by cysteine mapping, N-terminal sequencing, and tandem mass spectrometry analysis.

The protein composition of the crude venom of Sistrurus barbouri was analyzed by two-dimensional sodium dodecyl sulfate polyacrylamide gel electrophoresis. Proteins were separated by reversed phase high-performance liquid chromatography and characterized by N-terminal sequence analysis. The molecular mass and number of cysteine residues of the purified proteins were determined by matrix-associated laser desorption/ionization-time of flight mass spectrometry. Selected protein bands were subjected to in-gel tryptic digestion and peptide mass fingerprinting. Analysis of the tandem mass spectrometry spectra of selected doubly-charged peptide ions was done by collision-induced dissociation in a quadrupole-linear ion trap instrument. Our results show that the venom proteome of the pigmy rattlesnake S. barbouri is composed of proteins belonging to a few protein families, which can be structurally characterized by their disulfide bond contents.

Amino Acid Sequence↗
PubMed2003

Isolation of two novel mannan- and L-fucose-binding lectins from the green alga Enteromorpha prolifera: biochemical characterization of EPL-2.

EPL-1 and EPL-2 represent lectins isolated from the green alga Enteromorpha prolifera. Both lectins are 20- to 22-kDa single-chain, nonglycosylated proteins. N-terminal sequence analysis of peptides representing over 70% of their primary structures shows that EPL-1 and EPL-2 represent novel proteins. Sedimentation-diffusion equilibrium experiments showed that EPL-1 and EPL-2 had average apparent molecular masses of 60000+/-6000 Da (EPL-1) and 59500+/-3000 Da (EPL-2), indicating that EPL-1 and EPL-2 have a tendency to self-associate into higher order aggregates, possibly homodimers and homotetramers, in equilibrium. The carbohydrate-binding specificity of EPL-2 was studied by enzyme-linked lectin assay and intrinsic fluorescence measurements. The results show that the combining site of EPL-2 is capable of accommodating both D-mannose and L-fucose, which share the conformation of the hydroxyl groups at positions 2 (axial) and 4 (equatorial), and includes subsites for the substituents at O1 and for branched mannose residues.

Amino Acid Sequence↗
PubMed2003

Influence of porcine spermadhesins on the susceptibility of boar spermatozoa to high dilution.

The effect of heparin-binding and non-heparin-binding spermadhesins on the viability, motility, and mitochondrial activity of boar spermatozoa at the high dilution (300,000 sperm/ml) to which sperm are exposed during the process of sex sorting by flow cytometry was investigated. Incubation of spermatozoa with heparin-binding spermadhesins caused a time- and dose-dependent decrease in the percentage of functional spermatozoa. The percentage of viable spermatozoa incubated at 38 degrees C with heparin-binding spermadhesins diluted in PBS (1 mg/ml) dropped from 75% (0.5 h) to 4% (5 h), whereas the percentage of viable spermatozoa incubated in PBS without proteins (control) decreased from 85% (0.5 h) to 19% (5 h). Addition of non-heparin-binding PSP-I/PSP-II spermadhesin to the PBS resulted in a concentration-dependent increment of the percentage of viable cells (65% after 5-h incubation), with maximum effect at 1.5 mg/ml. The heparin-binding spermadhesins totally suppressed sperm motility and mitochondrial activity after 5 h of incubation. The same parameters of sperm incubated in the presence of 1.5 mg/ml of PSP-I/PSP-II were 50% and 58%, respectively, and the percentages of control sperm displaying motility and mitochondrial activity were 21% and 26%, respectively. Moreover, the viability, motility, and mitochondrial activity all decreased on incubation of spermatozoa with mixtures of PSP-I/PSP-II and heparin-binding spermadhesins as the concentration of the latter increased. We conclude that PSP-I/PSP-II and the heparin-binding spermadhesins exert antagonistic effects on the functionality of highly diluted boar spermatozoa. The finding that PSP-I/PSP-II contributes to maintaining sperm with high viability, motility, and mitochondrial activity for at least 5 h at physiological temperature points to its potential use as an additive for sperm preservation, specifically of highly diluted, flow-sorted spermatozoa for sex preselection.

Animals↗
PubMed2002

Crystal structure of a prostate kallikrein isolated from stallion seminal plasma: a homologue of human PSA.

Prostate-specific kallikrein, a member of the gene family of serine proteases, was initially discovered in semen and is the most useful serum marker for prostate cancer diagnosis and prognosis. We report the crystal structure at 1.42A resolution of horse prostate kallikrein (HPK). This is the first structure of a serine protease purified from seminal plasma. HPK shares extensive sequence homology with human prostate-specific antigen (PSA), including a predicted chymotrypsin-like specificity, as suggested by the presence of a serine residue at position S1 of the specificity pocket. In contrast to other kallikreins, HPK shows a structurally distinct specificity pocket. Its entrance is blocked by the kallikrein loop, suggesting a possible protective or substrate-selective role for this loop. The HPK structure seems to be in an inactivated state and further processing might be required to allow the binding of substrate molecules. Crystal soaking experiments revealed a binding site for Zn(2+) and Hg(2+), two known PSA inhibitors.

Amino Acid Sequence↗
PubMed2002

Sperm coating mechanism from the 1.8 A crystal structure of PDC-109-phosphorylcholine complex.

Bovine seminal plasma PDC-109 binds to sperm surface choline lipids and promotes sperm capacitation by stimulating the efflux of cholesterol and phospholipids. The structure of PDC-109 with bound phosphorylcholine was solved using MAD data of a single platinum site. Its two globular (40 x 50 x 20 A(3)) Fn2 domains are linked and clustered by a short polypeptide. The choline binding sites lie at the same face of the molecule. Phosphorylcholine binds to the Fn2 domains through a cation-pi interaction between the quaternary ammonium group and a core tryptophan, plus hydrogen bonding between hydroxyls of exposed tyrosines and the phosphate group. The structure of the PDC-109-oPC complex provides a structural ground for the sperm membrane-coating mechanism underlying PDC-109-induced capacitation.

Amino Acid Sequence↗
PubMedDate not supplied

Does seminal plasma PSP-I/PSP-II spermadhesin modulate the ability of boar spermatozoa to penetrate homologous oocytes in vitro?

Low concentration (0.15 mg per million of spermatozoa) of seminal plasma-derived PSP-I/PSP-II spermadhesin heterodimer is able to preserve the viability of highly extended boar spermatozoa. Whether spermatozoa also keep their fertilizing capacity is not yet known. The present study evaluated the effect of exposing freshly extended and frozen-thawed boar spermatozoa (10 million/mL) to PSP-I/PSP-II (1.5 mg/mL) for 30 or 120 minutes on sperm characteristics and the outcome of in vitro penetration of immature (IM) and in vitro matured (IVM) homologous oocytes, aiming to identify this spermadhesin as a suitable modulator for sperm-handling protocols. Although exposure to the heterodimer improved sperm viability and motility without increasing the levels of sperm acrosome exocytosis in both freshly extended and frozen-thawed spermatozoa, this pretreatment did not affect sperm penetration rates or sperm numbers per oocyte when pretreated fresh spermatozoa were coincubated with IM or IVM oocytes compared with controls. When cryopreserved spermatozoa were tested, however, on IVM oocytes, already a 30-minute preincubation exposure to PSP-I/PSP-II showed a significant blocking effect on penetration rate (from 90% to 32%, P < .05) and on mean sperm numbers per oocyte (2.9 to 1.6, P < .05). To disclose the nature of this paradox, frozen-thawed spermatozoa were cleansed (by centrifugation in saline bovine serum albumin or through Percoll density gradient separation) and the procedure repeated. Oocyte penetration (but not number of spermatozoa per oocyte) increased (P < .05) when spermatozoa were cleansed with Percoll compared with either washed or unwashed controls (53% vs 13% vs 31%, respectively). In addition, the percentages of polyspermic oocytes remained lower than control (38.5% vs 68.7%, respectively; P < .05). In conclusion, the results confirm that exposure of fresh or frozen-thawed boar spermatozoa to a low dose of seminal PSP-I/PSP-II spermadhesin preserves sperm viability and motility in vitro. Although there was no obvious influence of the heterodimer on the capability of freshly extended boar spermatozoa to penetrate homologous oocytes (either IM or IVM), PSP-I/PSP-II exerted a deleterious effect when frozen-thawed spermatozoa were used to penetrate IVM oocytes. Such an effect of cryopreservation seems to a certain extent reversible, since cleansing of the sperm surface decreased, at least partially, this blocking effect, increasing both penetration and the monospermic rates.

Animals↗