Purification, molecular cloning and heterologous expression of a glutathione S-transferase involved in insecticide resistance from the rice brown planthopper, Nilaparvata lugens

2002 ◽  
Vol 362 (2) ◽  
pp. 329-337 ◽  
Author(s):  
John G. VONTAS ◽  
Graham J. SMALL ◽  
Dimitra C. NIKOU ◽  
Hilary RANSON ◽  
Janet HEMINGWAY
Keyword(s):  
Insecticide Resistance ◽  
Peroxidase Activity ◽  
Pyrethroid Resistance ◽  
Brown Planthopper ◽  
Resistance Mechanism ◽  
Amino Acid Identity ◽  
Nilaparvata Lugens ◽  
Northern Analysis ◽  
Affinity Column ◽  
Glutathione S Transferase

A novel glutathione S-transferase (GST)-based pyrethroid resistance mechanism was recently identified in Nilaparvata lugens [Vontas, Small and Hemingway (2001) Biochem. J. 357, 65–72]. To determine the nature of GSTs involved in conferring this resistance, the GSTs from resistant and susceptible strains of N. lugens were partially purified by anion exchange and affinity chromatography. The majority of peroxidase activity, previously correlated with resistance, was confined to the fraction that bound to the affinity column, which was considerably elevated in the resistant insects. A cDNA clone encoding a GST (nlgst1-1) —the first reported GST sequence from Hemiptera with up to 54% deduced amino-acid identity with other insect class I GSTs—was isolated from a pyrethroid-resistant strain. Northern analysis showed that nlgst1-1 was overexpressed in resistant insects. nlgst1-1 was expressed in Escherichia coli, purified and characterized. The ability of the recombinant protein to bind to the S-hexylglutathione affinity matrix, its substrate specificities and its immunological properties confirmed that this GST was one from the elevated subset of N. lugens GSTs. Peroxidase activity of the recombinant nlgst1-1 indicated that it had a role in resistance, through detoxification of lipid peroxidation products induced by pyrethroids. Southern analysis of genomic DNA from the resistant and susceptible strains indicated that GST-based insecticide resistance may be associated with gene amplification in N. lugens.

scholarly journals Comparison of the knockdown resistance locus (kdr) in Anopheles stephensi, An. arabiensis, and Culex pipiens s.l. suggests differing mechanisms of pyrethroid resistance in east Ethiopia

2020 ◽  
Author(s):  
Tamar E. Carter ◽  
Araya Gebresilassie ◽  
Shantoy Hansel ◽  
Lambodhar Damodaran ◽  
Callum Montgomery ◽  
...  
Keyword(s):  
Insecticide Resistance ◽  
Culex Pipiens ◽  
Anopheles Stephensi ◽  
Pyrethroid Resistance ◽  
Resistance Mechanism ◽  
Target Site ◽  
Resistance Locus ◽  
Knockdown Resistance ◽  
Resistance Mutations ◽  
Vector Species

AbstractThe malaria vector, Anopheles stephensi, which is typically restricted to South Asia and the Middle East, was recently detected in the Horn of Africa. Controlling the spread of this vector could involve integrated vector control that considers the status of insecticide resistance of multiple vector species in the region. Previous reports indicate that the knockdown resistance mutations (kdr) in the voltage-gated sodium channel (vgsc) are absent in both pyrethroid resistant and sensitive variants of An. stephensi in east Ethiopia but similar information on other vector species in the same areas is limited. In this study, kdr and the neighboring intron was analyzed in An. stephensi, An. arabiensis, and Culex pipiens s. l. collected in east Ethiopia between 2016 and 2017. Sequence analysis revealed that all of Cx. pipiens s.l. (n = 42) and 71.6% of the An. arabiensis (n=67) carried kdr L1014F known to confer target-site pyrethroid resistance. Intronic variation was only observed in An. stephensi (segregating sites = 6, haplotypes = 3) previously shown to have no kdr mutations. In addition, no evidence of non-neutral evolutionary processes was detected at the An. stephensi kdr intron which further supports target-site mechanism not being a major resistance mechanism in this An. stephensi population. Overall, these results suggest differences in evolved mechanisms of pyrethroid/DDT resistance in populations of vector species from the same region. Variation in insecticide resistance mechanisms in East Ethiopian mosquito vectors highlight possible species or population specific biological factors and distinct environmental exposures that shape their evolution.

scholarly journals Purification and characterization of hepatic glutathione S-transferases of rhesus monkeys. A family of enzymes similar to the human hepatic glutathione S-transferases

1988 ◽  
Vol 251 (1) ◽  
pp. 81-88 ◽  
Author(s):  
R M Hoesch ◽  
T D Boyer
Keyword(s):  
Peroxidase Activity ◽  
Rhesus Monkeys ◽  
Affinity Column ◽  
Terminal Sequence ◽  
Glutathione S Transferase ◽  
Substrate Specificities ◽  
High Affinity ◽  
Human Enzyme ◽  
Hepatic Glutathione ◽  
Glutathione S Transferases

Thirteen forms of glutathione S-transferase were purified from the livers of female rhesus monkeys (Macaque mulatta). Most (74.7%) of the activity in the hepatic cytosol adhered well to the GSH affinity column and could be eluted only with the addition of GSH to the eluting buffer. The predominant isoenzymes (n = 5) in this ‘high-affinity’ fraction had alkaline pI values (greater than 9.0) and contained a subunit with an Mr value of 24,000. All of these isoenzymes had high organic peroxidase activity and, on the basis of amino acid analysis, substrate specificities and affinity for non-substrate ligands, appear to belong to the family of glutathione S-transferases that have been termed alpha [Mannervik, Alin, Guthenberg, Jensson, Tahir, Warholm & Jörnvall (1985) Proc. Natl. Acad. Sci. U.S.A. 82, 7202-7206]. Also within the high-affinity fraction was an isoenzyme with an acidic (5.8) pI value. This acidic isoenzyme was composed of a unique subunit (Mr 23,000). The N-terminal sequence (ten residues) of this acidic enzyme was identical with that of a human form that is referred to as pi. The predominant form of enzyme in the ‘low-affinity’ (eluted from the GSH affinity column with an increase in buffer pH) fraction was a homodimer of a 26,000-Mr subunit. It had an alkaline pI (greater than 9.0) but it lacked organic peroxidase activity. The N-terminal sequence (ten residues) of this enzyme was identical with that of a human enzyme referred to as mu. The substrate specificities and affinity for non-substrate ligands of this monkey enzyme also were similar to those of the human enzyme. In conclusion, the liver cytosol of rhesus monkeys contains a number of glutathione S-transferase isoenzymes that are very similar to the human hepatic enzymes.

Sign in / Sign up

Export Citation Format

Share Document