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At least 1,099 records · Page 61Linked to original sources
PubMed1993

Examination of the role of tyrosine-174 in the catalytic mechanism of the Arabidopsis thaliana chitinase: comparison of variant chitinases generated by site-directed mutagenesis and expressed in insect cells using baculovirus vectors.

Using the catalytic mechanism of lysozyme as a paradigm for the mechanism of other enzymes that catalyze the hydrolysis of beta-1,4-glycosidic linkages, including chitinase, we have examined the effect of chemical modification with 1-ethyl-3-(3-dimethylaminopropyl)-carbodiimide (EDC) on the reaction catalyzed by Zea mays chitinase. Inactivation with EDC did not result in derivatization of essential carboxylic acid residues, but resulted in the selective modification of a single essential tyrosine residue (Verburg, J. G., Smith, C. E., Lisek, C. A., and Huynh, Q. K., 1991, J. Biol. Chem. 267, 3886-3893). Here, we examine the role of the homologous tyrosine residue in the catalytic mechanism of the Arabidopsis thaliana chitinase. Tyrosine-174 of the Arabidopsis chitinase was replaced, with phenylalanine, alanine, histidine, and methionine by site-directed mutagenesis, and the variant chitinases were expressed in insect cells using baculovirus transfer vectors. A comparison of the reaction catalyzed by each of the variant enzymes indicates that substitution of another amino acid for Tyr-174 alters, but does not eliminate, enzymatic activity. Estimates of the specific activities of the variant chitinases reveal that substitution of His for Tyr-174 has a minimal effect on catalysis, the specific activities of the Phe and Met variants are approximately equivalent to each other, but are 60% the specific activity of wild-type Arabidopsis chitinase, and the specific activity of the Ala variant is only 40% that of wild-type. The observation that the Arabidopsis chitinase is tolerant to mutagenesis at this position suggests that Tyr-174 does not participate directly in catalysis.

Amino Acid Sequence↗
PubMed1990

Synthesis of foot-and-mouth disease virus capsid proteins in insect cells using baculovirus expression vectors.

Foot-and-mouth disease virus (FMDV) cDNA cassettes containing sequences encoding the capsid precursor P1-2A with and without those encoding the proteases L and 3C were introduced into Autographa californica nuclear polyhedrosis virus (AcMNPV) expression vectors. Procapsid proteins 1AB, 1C and 1D were produced in cells infected with recombinant baculoviruses, when L and 3C were present in the constructs, indicating that these FMDV proteases were active in insect cells. Unlike P1 processing in poliovirus, which has been shown to be catalysed mainly by the 3CD gene product, the 3C protease of FMDV was able to process P1 independently of 3D. Cytotoxicity of the L protease for insect cells prevented the use of the optimized transfer vector, pAcRP23, for inserting L-containing cassettes into AcMNPV. By contrast, viable AcMNPV-FMDV recombinants could be made without restriction on choice of the transfer vector when the L gene was either not expressed or inactivated by an inframe deletion. In the latter case, normal cleavage at the L-P1 junction no longer occurred in cis, and a new processing event, probably catalysed by 3C, was observed within the C-terminal region of the residual L protein. Analysis of baculovirus-expressed products in sucrose gradients showed that a fraction of the capsid proteins is present in an aggregated form, migrating at 70S and possibly resembling FMDV empty capsid particles.

Animals↗
PubMed1997

[Preliminary evaluation of the incidence and control of insects--pest control in Polish hospitals].

The evaluation of the infestation and methods of insect disease vectors control in 748 hospitals in Poland in the period of 1990 to 1995 were done. The insect species, places of their occurrence and control agents were analysed. Blattella germanica L. occurred most frequently (71% hospitals). Blatta orientalis and Monomorium pharaonis were found in 40% and 17% hospitals respectively. Kitchens, laundries and baths were most infested. Sometimes insects were found also in central sterilization units and operating theaters. Controls of insects in hospitals were performed one to four times a year mostly by spraying with residual formulation. The control agents contained pyrethroids (mostly permethrin, but also deltamethrin and cypermethrin) and carbamates (bendiocarb, propoxur). Baits with hydramethylnon, boric acid, methoprene and chlorpyrifos were used not very often. The authors suggest reduction in using the spraying agents. The baits are recommended because they delay the development of the resistance to pesticides in controlling insect populations and are safer.

Animals↗
PubMed2003

Post-integration behavior of a Mos1 mariner gene vector in Aedes aegypti.

The post-integration behavior of insect gene vectors will determine the types of applications for which they can be used. Transposon mutagenesis, enhancer trapping, and the use of transposable elements as genetic drive systems in insects requires transposable elements with high rates of remobilization in the presence of transposase. We investigated the post-integration behavior of the Mos1 mariner element in transgenic Aedes aegypti by examining both germ-line and somatic transpositions of a non-autonomous element in the presence of Mos1 transposase. Somatic transpositions were occasionally detected while germ-line transposition was only rarely observed. Only a single germ-line transposition event was recovered after screening 14,000 progeny. The observed patterns of transposition suggest that Mos1 movement takes place between the S phase and anaphase. The data reported here indicate that Mos1 will be a useful vector in Ae. aegypti for applications requiring a very high degree of vector stability but will have limited use in the construction of genetic drive, enhancer trap, or transposon tagging systems in this species.

Aedes↗