Crystals of the Chlamydomonas reinhardtii cell wall: polymerization, depolymerization, and purification of glycoprotein monomers

Two of the three major outer layers of the Chlamydomonas reinhardtii cell wall (W6 and W4) can be solubilized from living cells with sodium perchlorate or other chaotropes and will repolymerize in vitro to form milligram amounts of wall crystals. Conditions for optimal crystalization are presented, and conditions that fail to induce polymerization are exploited to maintain monomers in aqueous solution for ion-exchange chromatography. The four major glycoproteins of the complex (GP1, 1.5, 2, and 3) have in this way been purified to apparent homogeneity and have been characterized morphologically by transmission electron microscopy using the quick-freeze, deep-etch technique and by amino acid composition. Three of the four are hydroxyproline-rich species that copolymerize to form the W6 layer. The fourth (GP1.5) is a glycine-rich species that binds to the interior of the in vitro crystal; it is apparently equivalent to the granules within the W4 layer in situ.     

Alterations to the catalytic properties of polyphenoloxidase in detergentless microemulsions and ternary water-organic solvent mixtures

Summary The catalytic properties of mushroom polyphenoloxidase could be substantially altered by entrapment into hexane- and toluene-based microemulsions stabilized with isopropanol. The fast irreversible inactivation and drastic substrate inhibition of the enzyme were significantly reduced in detergentless microemulsions in comparison to conventional aqueous media. Similar changes in the catalytic behavior of polyphenoloxidase were observed in the normal ternary solutions of hexane-(toluene)-isopropanol-water, and in the H-bonded aggregates of isopropanol and water in toluene, but not in hexane.     

The Comparative Wood-destroying Ability and Preservative Tolerance of Monocaryotic and Dicaryotic Mycelia of Lenzites trabea (Pers.) Fr. and Poria vaillantii (DC ex Fr.) Cke.

Comparison by soil-block tests on several timbers of the wood-destroyingabilities of dicaryotic cultures of both Poria vaillantii andLenzites trabea with those of monocaryotic cultures either derivedfrom, or contributing to, the formation of the dicaryon, indicatedthat monocaryotic cultures of P. vaillantii were generally moredestructive than related dicaryotic cultures, whereas dicaryonsof L. trabea tended to be slightly more destructive than relatedmonocaryons. The tolerance of L. trabea monocaryons to coppersulphate and to sodium arsenate in nutrient agar showed someindication of being higher than that of related dicaryons, whilstthe tolerance of P. vaillantii monocaryons to a copperchrome-arsenatepreservative in soil-block tests also appeared to be higherthan that of related dicaryons. It is concluded that, althoughneither wood-destroying ability nor preservative tolerance wasgreatly affected by nuclear condition, the differences shownmay be of importance in conjunction with the monocaryotizationof dicaryons by toxic agents during laboratory decay tests.     

High-pressure liquid chromatography fractionation of Chlamydomonas dynein extracts and characterization of inner-arm dynein subunits

A rapid procedure for fractionating salt-stable dynein subunits from high-salt extracts of Chlamydomonas axonemes has been developed using a high-pressure liquid chromatography system with an anion exchange column and gradient salt elution. Five distinct fractions are shown to be highly enriched for five distinct subunits or subunit complexes by SDS/polyacrylamide gel electrophoresis. ATPase activity and electron microscopy. Peaks 1 and 4 contain, respectively, the single-headed gamma-subunit and the two-headed alpha/beta-heteropolymer that form the outer arm in situ and are dissociated by salt exposure; both peaks are absent from the outer arm-less mutant pf-28. Peaks 2, 3 and 5 contain, respectively, two distinct single-headed species and a double-headed species that derive from inner arms; all three peaks are missing from the inner arm-less mutant pf-23. Sucrose-gradient sedimentation analysis confirms these assignments and provides additional information on the intermediate-chain and light-chain composition of the inner-arm species. Electron microscopy of the purified inner-arm species visualized by the quick-freeze deep-etch technique complements a previous analysis of outer-arm species. Each protein is shown to have a unique morphology, and both the inner- and outer-arm proteins clearly belong to a common family whose structural divergence presumably reflects functional specialization.     

Structural and functional differentiation of two clinally distributed glucosephosphate isomerase allelic isozymes from the teleost Fundulus heteroclitus

The teleost Fundulus heteroclitus (L.) possesses two loci, Gpi-A and Gpi-B, for the glycolytic enzyme, glucose-phosphate isomerase (GPI; D- glucose-6-phosphate ketol-isomerase; E.C. 5.3.1.9). The Gpi-B locus is polymorphic in Fundulus, with two common alleles, Gpi-Bb and Gpi-Bc, distributed in a clinal manner in populations along the east coast of North America. Since this clinal distribution is strongly correlated with a temperature gradient, we asked whether the GPI-B2 allozymes were functionally adapted to the thermal environment in which a given phenotype predominated. The two major GPI-B2 allozymes were purified to homogeneity and were characterized as to molecular weight, isoelectric pH, thermal denaturation, and kinetic parameters. Both GPI-Bb2 and GPI- Bc2 allozymes have molecular masses of 110 kD, and they have isoelectric pHs of 6.4 and 6.6, respectively. The GPI-Bb2 allozyme was more stable to thermal denaturation than was the GPI-Bc2 enzyme. Kinetic properties of the allelic isozymes were investigated both as a function of pH and as a function of temperature. At 25 degrees C, over the pH range considered, there were no significant differences between allozymes, either in Km for fructose-6-phosphate or in Ki for 6- phosphogluconate, but apparent Vmax values differed between pH 7.5 and pH 8.5. All steady-state kinetic parameters showed strong temperature dependence, but the allozymes differed only in the Ki for 6- phosphogluconate at temperatures greater than 30 degrees C. On the basis of the observed structural and functional differences alluded to above, the hypothesis that the major allelic isozymes of the Gpi-B locus were functionally equivalent was rejected. However, it is not yet known whether these structural and functional differences have any significance at higher levels of biological organization.      

Can Handheld Thermal Imaging Technology Improve Detection of Poachers in African Bushveldt?

Illegal hunting (poaching) is a global threat to wildlife. Anti-poaching initiatives are making increasing use of technology, such as infrared thermography (IRT), to support traditional foot and vehicle patrols. To date, the effectiveness of IRT for poacher location has not been tested under field conditions, where thermal signatures are often complex. Here, we test the hypothesis that IRT will increase the distance over which a poacher hiding in African scrub bushveldt can be detected relative to a conventional flashlight. We also test whether any increase in effectiveness is related to the cost and complexity of the equipment by comparing comparatively expensive (22000 USD) and relatively inexpensive (2000 USD) IRT devices. To test these hypotheses we employ a controlled, fully randomised, double-blind procedure to find a poacher in nocturnal field conditions in African bushveldt. Each of our 27 volunteer observers walked three times along a pathway using one detection technology on each pass in randomised order. They searched a prescribed search area of bushveldt within which the target was hiding. Hiding locations were pre-determined, randomised, and changed with each pass. Distances of first detection and positive detection were noted. All technologies could be used to detect the target. Average first detection distance for flashlight was 37.3m, improving by 19.8m to 57.1m using LIRT and by a further 11.2m to 68.3m using HIRT. Although detection distances were significantly greater for both IRTs compared to flashlight, there was no significant difference between LIRT and HIRT. False detection rates were low and there was no significant association between technology and accuracy of detection. Although IRT technology should ideally be tested in the specific environment intended before significant investment is made, we conclude that IRT technology is promising for anti-poaching patrols and that for this purpose low cost IRT units are as effective as units ten times more expensive.     

Phylogenetic relationships among marsupial frog genera (Anura: Hylidae: Hemiphractinae) based on evidence from morphology and natural history

We reassess the phylogenetic relationships of genera of hemiphractine hylid frogs (Marsupial Treefrogs) and discuss the evolution of several distinctive characters within this group using parsimony analysis. Fifty-one morphological and life-history characters were sampled from two species of Cryptobatrachus , three species of Flectonotus , 17 species of Gastrotheca , all five species of Hemiphractus , and one species of Stefania as the ingroup and three hyline, one phyllomedusine, and one pelodryadine species as outgroups. Our results support the mon-ophyly of Flectonotus, Cryptobatrachus , and Hemiphractus. Gastrotheca is paraphyletic with respect to Hemiphractus , dorsal pouches were lost in the ancestor of Hemiphractus. Direct development is a synapomorphy for Hemiphractinae and tadpoles were regained independently several times. These results stand in stark contrast to the prevailing paradigm regarding marsupial frog relationships.     

The role of the blood meal in egg-laying periodicity and fecundity in Triatoma infestans

The number of eggs produced by Triatoma infestans is determined by the amount of blood ingested; the formation of one egg requires 16 mg blood. An exogenous supply of juvenile hormone, provided by the implantation of extra corpora aliata, increases egg production per unit of blood ingested. The corpus allatum is therefore implicated in the relationship between nutrition and fecundity. The rhythm of egg production is regulated by the nutritional state of the adult female: when the females are fed at frequent intervals, egg production becomes regular and continuous. This calls into question previous interpretations of egg-laying rhythms in the Triatominae as cyclic.