Latex Collection Research Articles

Purification and characterization of a new chitinase from latex of Ipomoea carnea

A novel enzyme with endochitinase/lysozyme activity was purified to homogeneity from latex of Ipomoea carnea subsp. fistulosa using latex collection, gum removal, ammonium sulphate precipitation, hydrophobic interaction, and anion exchange chromatography. The enzyme was glycosylated (5–6%) and homogeneous on SDS-PAGE; has a molecular mass of 30.06 kDa (MALDI-TOF) and an isoelectric point of pH 4.6. The enzyme exhibited chitinase activity for hydrolysis of glycol chitin and the chitinolytic activity was significantly inhibited by allosamidin and mercuric chloride. The enzyme is stable in the pH range of 4.0–9.5, 80 °C and the optimal activity was observed at pH 5.5 and 50 °C. The enzyme consists of 8 tryptophan, 14 tyrosine, 6 cysteine residues forming three disulfide bridges and the extinction coefficient was estimated as 21.35 M −1 cm −1. The polyclonal antibody was raised in rabbit and immunodiffusion suggests that the antigenic determinants are unique. The first 15 N-terminal residues G-E-I-T-I-Y-W-G-Q-N-G-F-E-G-S exhibited high identity to other known plant chitinases. Owing to the economic purification, high yield, unique and extraordinary features, stability and behavior; the enzyme ICChII can be widely employed in agriculture, industry, environmental protection, and in recycling chitinous waste from arthropod shellfish and for chito-oligosaccharide production.

A convenient and efficient protocol for isolating high-quality RNA from latex of Hevea brasiliensis (para rubber tree)

Isolating high-quality RNA from latex of H. brasiliensis is a prerequisite to elucidating the molecular mechanisms of rubber biosynthesis and its regulation. Here, an improved protocol was developed for latex collection, transportation, storage, and RNA isolation. Compared with existing ones, our protocol eliminated liquid nitrogen for latex collection and subsequent low-temperature (− 70 °C) condition for latex storage, making it more convenient and feasible when latex was collected in remote sampling sites, and latex storage and RNA isolation were conducted in poorly-equipped laboratories. Different methods (UV absorbance scans, denaturing gel electrophoresis, autoradiograph monitoring of cDNA synthesis) were used to confirm the high quality of the RNA prepared with this protocol, whose usefulness was further verified by several practical applications, including construction of one high-quality cDNA library, cloning of the full-length cDNAs of 3 novel Hevea sucrose transporter genes, and semi-quantitative RT-PCR analysis of two rubber-biosynthesis essential genes and one sucrose transporter gene.

Isolation and characterization of isoinhibitors of the potato protease inhibitor I family from the latex of the rubber trees, Hevea brasiliensis

Three isoinhibitors have been isolated to homogeneity from the C-serum of the latex of the rubber tree, Hevea brasiliensis clone RRIM 600, and named HPI-1, HPI-2a and HPI-2b. The three inhibitors share the same amino acid sequence (69 residues) but the masses of the three forms were determined to be 14,893 ± 10, 7757 ± 5, and 7565 ± 5, respectively, indicating that post-translational modifications of the protein have occurred during latex collection. One adduct could be removed by reducing agents, and was determined to be glutathione, while the other adduct could not be removed by reducing agents and has not been identified. The N-termini of the inhibitor proteins were blocked by an acetylated Ala, but the complete amino acid sequence analysis of the deblocked inhibitors by Edman degradation of fragments from endopeptidase C digestion and mass spectrometry confirmed that the three isoinhibitors were derived from a single protein. The amino acid sequence of the protein differed at two positions from the sequence deduced from a cDNA reported in GenBank. The gene coding for the inhibitor is wound-inducible and is a member of the potato inhibitor I family of protease inhibitors. The inhibitor strongly inhibited subtilisin A, weakly inhibited trypsin, and did not inhibit chymotrypsin. The amino acid residues at the reactive site P 1 and P 1 ′ were determined to be Gln45 and Asp46, respectively, residues rarely reported at the reactive site in potato inhibitor I family members. Comparison of amino acid sequences revealed that the HPI isoinhibitors shared from 33% to 55% identity (50–74% similarity) to inhibitors of the potato inhibitor I family. The properties of the isoinhibitors suggest that they may play a defensive role in the latex against pathogens and/or herbivores.

Detection of three wound-induced proteins in papaya latex

The effects of routine mechanical wounding for latex collection from unripe fruits of the tropical Carica papaya tree were investigated. For that purpose, the protein composition of three different latexes was analyzed. The first one, commercially available, was provided in the form of a spray-dried powder, the second one was harvested from fully grown but unripe papaya fruits that are regularly tapped for latex production and the last one, was obtained from similar fruits wounded for the first time. Repeated mechanical wounding was found to profoundly affect the protein content of the latex inducing, among others, activation of papain. Regularly tapped latexes also accumulated several low molecular weight proteins not yet identified, as well as three proteins identified as a trypsin inhibitor, a class-II chitinase and a glutaminyl cyclase on the basis of their enzymatic or inhibitory activities and chromatographic elution profiles. This latter was found here, for the first time, to be a wound-induced protein. The roles of these proteins in the plant defense mechanism are discussed.

Allergenic proteins of natural rubber latex

As the living cytoplasm of laticiferous cells, Hevea brasiliensis latex is a rich blend of organic substances that include a mélange of proteins. A small number of these proteins have given rise to the problem of latex allergy. The salient characteristics of H. brasiliensis latex allergens that are recognized by the International Union of Immunological Societies (IUIS) are reviewed. These are the proteins associated with the rubber particles, the cytosolic C-serum proteins and the B-serum proteins that originate mainly from the lutoids. Procedures for the isolation and purification of latex allergens are discussed, from latex collection in the field to various preparative approaches adopted in the laboratory. As interest in recombinant latex allergens increases, there is a need to validate recombinant proteins to ascertain equivalence with their native counterparts when used in immunological studies, diagnostics, and immunotherapy.

The dependence of crude papain yields on different collection (‘tapping’) procedures for papaya latex

AbstractThe yields of fresh papaya latex, dry latex (crude papain) and proteolytic activity were studied with respect to fruit age (1.5–3 months); sex of the tree (those which produce fruit: female and hermaphrodite); time (5:30 a.m.‐2:00 p.m.) of latex collection; pattern and frequency of fruit incisions. The yields from 250 randomly selected trees (750 fruits) were investigated under these different conditions. Yields of fresh latex are frequently unreliable guides to yields of proteolytic activity. In contrast to the literature recommendations, the yields of crude papain from female and hermaphrodite trees and during different periods of the day were similar. The proteolytic activity differs significantly with time of collection only with hermaphrodite trees. The proteolytic activity of hermaphrodite crude papain is usually less than that from females (under some conditions only 60% of that from females). Yields of crude papain were higher in older fruits (2.5–3 months) from both sexes. The proteolytic activity of the crude papain was highest in the younger fruits (1.5–2 months) from female trees (30% higher than from older fruits); these activity differences with age were not significant with hermaphrodite trees. No significant differences in crude papain yields were encountered using three or six incisions per fruit at weekly intervals. The optimum procedure is to tap 2.5–3‐month‐old fruit of both sexes with three incisions between dawn and the start of rains in early afternoon, at about 4‐day intervals.

Method of Testing the Mechanical Properties of Vulcanizates Prepared from Latex as a Starting Material

Abstract With the method which has been described, it is possible to obtain reproducible results easily. It is hoped that the method will make it easier to study the properties of rubber mixtures prepared from latex. Furthermore the quantity of a latex mixture needed for such a test is small, i.e., of the order of 400 cc., so the method is particularly well adapted to the selection of Hevea trees on a basis of the quality of their latex. As a matter of fact, it will be easy in many cases to follow the collection of latex from a limited number of trees. It should even be possible, by using a single dipping form and reducing the size of the dipping tank, to follow the quality of the latex from the daily production of one tree, a procedure which is impossible with coagulated rubber.