Proteinase activity in the white shrimp (Penaeus vannamei) clotting protein.

The clottable protein (CP) was isolated from white shrimp, Penaeus vannamei plasma as a 400-kDa protein that splits to two identical 200-kDa subunits when it is reduced with 2-ME. However, using DTT as reducing agent, four main bands were observed; two of them (179 and 125 kDa) had the same N-terminus sequence of the intact CP, indicating that most fragmentation occurs in the carboxy-terminus. The proteinase activity of reduced CP was detected using azoalbumin as substrate. Proteinase activity was only detected in the reduced, but not alkylated protein. Trypsin and papain, as well as soybean trypsin inhibitor and E64, were included for comparison. Proteolytic activity of reduced CP was inhibited by E64, but not by STI, indicating that such activity corresponds to a cysteine type proteinase.

Prophenoloxidase from brown shrimp (Penaeus californiensis) hemocytes.

Prophenoloxidase (proPO) was purified from blood cells of the brown shrimp Penaeus californiensis by ultracentrifugation and dye affinity chromatography. The isolated proPO is a 114-kDa monomeric protein as determined by SDS-PAGE. This protein can be hydrolyzed by proteinases, producing a 107-kDa active phenoloxidase (PO). The isoelectric point for both protein forms was 7.35. The PO reaction using L-DOPA as substrate, has an optimum pH of 8, and was poorly inhibited by sodium azide, thiourea and EDTA, but strongly inhibited by diethyl thiocarbamate. According to the substrate affinity and inhibition characteristics, this phenoloxidase was classified as a tyrosinase-like phenoloxidase. Purified proPO was not activated by bacterial lipopolysaccharides or beta-glucans.

Stability of some Cactaceae proteins based on fluorescence, circular dichroism, and differential scanning calorimetry measurements.

Characterization of three cactus proteins (native and denatured) from Machaerocereus gummosus (Pitahaya agria), Lophocereu schottii (Garambullo), and Cholla opuntia (Cholla), was based on electrophoretic, fluorescence, CD (circular dichroism), DSC (differential scanning calorimetry), and FT-IR (Fourier transform infrared) measurements. The obtained results of intrinsic fluorescence, DSC, and CD were dissimilar for the three species of cactus, providing evidence of differences in secondary and tertiary structures. Cactus proteins may be situated in the following order corresponding to their relative stability: Machaerocereus gummosus (Pitahaya agria) > Cholla opuntia (Cholla) > Lophocereu schottii (Garambullo). Thermodynamic properties of proteins and their changes upon denaturation (temperature of denaturation, enthalphy, and the number of ruptured hydrogen bonds) were correlated with the secondary structure of proteins and disappearance of alpha-helix.

Shrimp plasma HDL and beta-glucan binding protein (BGBP): comparison of biochemical characteristics.

A high density lipoprotein (HDL) and beta-glucan binding protein (BGBP) have been found in the hemolymph of marine shrimp. These proteins are involved in the transport of lipid and the recognition of foreign matter, respectively. Similarities in the color of the proteins and the molecular mass were noted. For a detailed comparison, HDL and BGBP were purified from two shrimp species, Penaeus vannamei and Penaeus californiensis, and their biochemical characteristics determined. Both proteins from each of the shrimp species are monomeric with approximately the same molecular mass in sodium dodecyl sulfate-polyacrylamide gel electrophoresis (approximately 100-112 kDa) and contain carbohydrate and lipid. The amino acid composition is similar and there is a high degree of similarity in the N-terminus. Furthermore, they are recognized by antibodies prepared independently. These results reveal that BGBP and HDL in shrimp hemolymph are the same protein, suggesting that there is a close relationship between the ability to respond to foreign matter and the diet as a provider of essential nutrients.

Isolation of Vibrio and Pseudomonas from brown shrimp (Penaeus californiensis Holmes) intestine.

Bacteria of the genera Vibrio, Pseudomonas and Aeromonas were isolated from the intestine of apparently healthy brown shrimp (Penaeus californiensis Holmes) cultured in a tidal pond. Species from these genera of bacteria have been reported as shrimp pathogens and have been involved in human gastrointestinal disorders related to seafood consumption. Isolation was done first in Marine broth, then in selective media (TCBS, Cetrimide and MacConkey). The oxidase negative strains were discarded as insignificant to shrimp culture. The identification of oxidase positive strains was based in morphological and colonial characteristics, biochemical capabilities, and both salinity and temperature tolerance. API 20E system and fatty acid analysis were also included. Three potentially pathogenic bacteria, Vibrio parahaemolyticus, Vibrio furnissii and Pseudomonas putida were isolated and identified from healthy shrimp intestine.

Purification and comparison of beta-1,3-glucan binding protein from white shrimp (Penaeus vannamei).

A beta-glucan binding protein (BGBP) was identified in both white (Penaeus vannamei) and blue shrimp (P. stylirostris) plasma. White shrimp BGBP was purified by affinity chromatography using immobilized laminarin, and its molecular and biological properties were described. White shrimp BGBP is a monomeric protein with a molecular mass of 100 kDa, similar to those described for other crustacean BGBPs. White and blue shrimp BGBPs can be detected with antisera against crayfish BGBP and brown shrimp BGBP. Both amino acid composition and N-terminal sequence are markedly similar to brown shrimp (P. californiensis) and crayfish (Pacifastacus leniusculus) BGBP, indicating that this recognition protein is present in freshwater and marine crustaceans.

A plasma protein isolated from brown shrimp (Penaeus californiensis) which enhances the activation of prophenoloxidase system by beta-1,3-glucan.

A beta-glucan-binding protein (BGBP) has been identified in brown shrimp plasma by using a polyclonal antiserum against a BGBP from the freshwater crayfish. The protein was purified by immunoaffinity chromatography, and its molecular and biological properties described. Brown shrimp BGBP is a monomeric protein with a molecular mass of 100 kDa, similar to those described for other crustacean BGBPs. This protein is capable of enhancing the prophenoloxidase (proPO) system activation induced by laminarin. Both amino acid composition and N-terminal sequence are markedly similar to those of crayfish BGBP.

Isolation of an immunosuppressive lectin from Phaseolus vulgaris L. cv Cacahuate using stroma.

An immunosuppressive lectin was isolated from seed of Phaseolus vulgaris cv Cacahuate using physically entrapped stroma. The lectin was found to be a 94 kDa tetrameric protein. When 50 micrograms, of this lectin were administered intraperitoneally 2 days before the immunization with sheep red blood cells, humoral response against the immunogen was completely inhibited. Other properties of the protein are discussed.

Purification and characterization of a lectin from Phaseolus acutifolius var. latifolius.

The lectin from Phaseolus acutifolius var. latifolius has been purified to homogeneity by affinity chromatography using stroma-sephadex, gel filtration and electrofocusing. The purified lectin consists of four subunits of 21 Kd molecular mass each one with 7.4% w/w total carbohydrate. Red cells of human and animal species, are agglutinated but are not inhibited by a series of sugars. Mitogenicity and immunosuppressive activities were demonstrated. Agglutination requires magnesium and calcium ions.