Structural basis of hAT transposon end recognition by Hermes, an octameric DNA transposase from Musca domestica.

Hermes is a member of the hAT transposon superfamily that has active representatives, including McClintock's archetypal Ac mobile genetic element, in many eukaryotic species. The crystal structure of the Hermes transposase-DNA complex reveals that Hermes forms an octameric ring organized as a tetramer of dimers. Although isolated dimers are active in vitro for all the chemical steps of transposition, only octamers are active in vivo. The octamer can provide not only multiple specific DNA-binding domains to recognize repeated subterminal sequences within the transposon ends, which are important for activity, but also multiple nonspecific DNA binding surfaces for target capture. The unusual assembly explains the basis of bipartite DNA recognition at hAT transposon ends, provides a rationale for transposon end asymmetry, and suggests how the avidity provided by multiple sites of interaction could allow a transposase to locate its transposon ends amidst a sea of chromosomal DNA.

Functional characterization of CYP6G4 from the house fly in propoxur metabolism and resistance.

The house fly (Musca domestica L.) (Diptera: Muscidae) is a global vector that can transmit >250 human and animal diseases. The control of house flies has heavily relied on the application of various chemical insecticides. The carbamate insecticide propoxur has been widely used for the control of house flies, and resistance to propoxur has been documented in many house fly populations worldwide. Previous studies have identified several propoxur resistance-conferring mutations in the target protein acetylcholinesterase; however, the molecular basis for metabolic resistance to propoxur remains unknown. In this study, we investigated the involvement of CYP6G4, a cytochrome P450 overexpressed in many insecticide resistant populations of Musca domestica, in propoxur metabolism and resistance by using combined approaches of recombinant protein-based insecticide metabolism and the Drosophila GAL4/UAS transgenic system. The recombinant CYP6G4 and its redox partners (NADPH-dependent cytochrome P450 reductase and cytochrome b5) were functionally expressed in Escherichia coli. Metabolism experiments showed that CYP6G4 was able to transform propoxur with a turnover rate of around 0.79 min-1. Six metabolites were putatively identified, suggesting that CYP6G4 could metabolize propoxur via hydroxylation, O-depropylation and N-demethylation. Moreover, bioassay results showed that ectopic overexpression of CYP6G4 in fruit flies significantly increased their tolerance to propoxur. Our in vivo and in vitro data convincingly demonstrate that CYP6G4 contributes to propoxur metabolism and resistance.

Midgut fluxes and digestive enzyme recycling in Musca domestica: A molecular approach.

Insects are reported to have water midgut countercurrents fluxes powering enzyme recovery before excretion, usually known as enzyme recycling. Up to now there is a single, and very incomplete, attempt to relate transporters and channels with countercurrent fluxes. In this work, M. domestica midgut water fluxes were inferred from the concentration of ingested and non absorbable dye along the midgut, which anterior midgut was divided in two sections (A1, A2), the middle in one (M) and the posterior midgut in four (P1, P2, P3, and P4), which led to the finding of additional sites of secretion and absorption. Water is secreted in A1 and A2 and absorbed at the middle midgut (M), whereas in posterior midgut, water is absorbed at P2 and secreted in the other sections, mainly at P4. Thus, a countercurrent flux is formed from P4 to P2. To disclose the involvement of the known water transporters Na+:K+:2Cl- (NKCC) and K+:Cl- (KCC), as well as the water channels aquaporins in water fluxes, their expression was evaluated by RNA-seq analyses from triplicate samples of seven sections along the midgut. MdNKCC1 was expressed in A1, MdNKCC2 was expressed in M1 and P2 and MdKCC in middle and in the most posterior region, thus apparently involved in secretion, absorption and both, respectively. MdNKCC2, MdKCC and aquaporins MdDRIP1 and 2 were confirmed as being apical by proteomics of purified microvillar membranes. The role of NKCC and KCC on midgut water fluxes was tested observing the effect of the inhibitor furosemide. The change of trypsin distribution along the posterior midgut and the increase of trypsin excretion in the presence of furosemide lend support to the proposal that countercurrent fluxes power enzyme recycling and that the fluxes are caused by NKCC and KCC transporters helped by aquaporins.

Design, Synthesis and Insecticide Activity of Novel Acetylcholinesterase Inhibitors: Triazolinone and Phthalimide Heterodimers.

Based on the "cluster effect" and the structure characters of acetylcholinesterase (AChE; EC 3.1.1.7), a new series of 1,2,4-triazolin-3-one and phthalimide heterodimers were designed, synthesized, and evaluated as potent dual acetylcholinesterase inhibitors (AChEIs). Most of the synthesized compounds showed good in vitro inhibitory activities towards both Drosophila melanogaster acetylcholinesterase (DmAChE) and Musca domestica acetylcholinesterase (MdAChE). Among them, 5g was found to be the most potent anti-AChE derivate (5g, IC50 = 8.07 µM to DmAChE, IC50 = 32.24 µM to MdAChE). It was 2.31- and 1.35-fold more active than the positive control ethion (CP, IC50 = 18.62 µM to DmAChE, IC50 = 43.56 µM to MdAChE). The docking model study revealed that 5g possessed the fitted spatial structure and bound to the central pocket and peripheral site of DmAChE. Moreover, most compounds demonstrated high insecticidal activity to Lipaphis erysimi and Tetranychus cinnabarinus at the concentration of 300 mg/L.

Crystal structure of the delta-class glutathione transferase in Musca domestica.

Among the various glutathione transferase (GST) isozymes in insects, the delta- and epsilon-class GSTs fulfill critical functions during the detoxification of insecticides. We crystalized MdGSTD1, the major delta-class GST isozyme in the housefly (Musca domestica), in complex with glutathione (GSH) and solved its structure at a resolution of 1.4 Å. The overall folding of MdGSTD1 resembled other known delta-class GSTs. Its substrate binding pocket was exposed to solvent and considerably more open than in the epsilon-class GST from M. domestica (MdGSTE2). However, their C-terminal structures differed the most because of the different lengths of the C-terminal regions. Although this region does not seem to directly interact with substrates, its deletion reduced the enzymatic activity by more than 70%, indicating a function in maintaining the proper conformation of the binding pocket. Binding of GSH to the GSH-binding region of MdGSTD1 results in a rigid conformation of this region. Although MdGSTD1 has a higher affinity for GSH than the epsilon class enzymes, the thiol group of the GSH molecule was not close enough to serine residue 9 to form a hydrogen-bond with this residue, which is predicted to act as the catalytic center for thiol group deprotonation in GSH.

Characterization of a new multifunctional beta-glucosidase from Musca domestica.

OBJECTIVE: To engineer Pichia pastoris for heterologous production of cellulase from Musca domestica and explore its potential for industrial applications. RESULTS: A new beta-glucosidase gene (bg), encoding 562 amino acids, was cloned from M. domestica by using rapid amplification of cDNA ends. The gene bg was linked to pPICZαA and expressed in P. pastoris with a yield of 500 mg l-1. The enzyme has the maximum activity with 27.6 U mg-1 towards cellulose. The beta-glucosidase has stable activity from 20 to 70 °C and can tolerate one-mole glucose. It has the maximum activities for salicin (25.9 ± 1.8 U mg-1), cellobiose (40.1 ± 2.3 U mg-1) and cellulose (27.6 ± 3.5 U mg-1). The wide-range substrate activities of the beta-glucosidase were further verified by matrix-assisted laser desorption/ionization mass spectra. Structural analysis shows that the beta-glucosidase belongs to glycoside hydrolase family Ι and possesses O-glycosylation sites. CONCLUSIONS: Thus, a multifunctional beta-glucosidase was expressed from M. domestica and provides a potential tool for industrial application of cellulose.

Binary combinations of organophosphorus pesticides exhibit differential toxicity under oxidised and un-oxidised conditions.

Acetylcholinesterase (AChE) inhibition has been demonstrated to be useful as a biomarker for exposure to organophosphorus (OP) insecticides in many environments. The objective of this study was to investigate the response of housefly (Musca domestica) head AChE (HF-AChE) exposed to five OPs as individual compounds and their binary mixtures under in vitro conditions. To examine the effects of oxidation on OP potency in the HF-AChE system, bromine water was used as an oxidisng agent. With oxidation, the sensitivity of HF-AChE to chlorpyrifos (CPF), malathion (MLT) and triazophos (TRZ) increased significantly. Monocrotophos (MCP) and profenofos (PRF) did not exhibit any significant differences in toxicity under oxidised and un-oxidised conditions. The toxicological interaction of five organophosphorus pesticides was evaluated using the concentration addition model, the combination index-isobologram equation and the toxic unit approach. All three models provided similar predictions for the 10 binary combinations of OPs under oxidised and un-oxidised conditions. In the present study, the antagonistic effects of the binary combination of OPs (CPF+PRF, CPF+MLT, MCP+MLT, PRF+MLT, MLT+TRZ and PRF+TRZ) were observed under oxidised conditions. This may be due to dispositional and/or receptor antagonism. Most of the binary combinations assayed under un-oxidised conditions exhibited synergistic responses. Triazophos showed very strong synergism in binary combinations with CPF, MCP and PRF un-oxidised conditions. In contrast, under oxidised conditions, only CPF+TRZ exhibited synergism. The results obtained indicate differential toxicity of binary combinations of OPs under oxidised and un-oxidised conditions. This information could be a valuable tool in understanding the mechanisms of OPs interactions and the interpretation of future in vivo studies with mixtures of OP insecticides.

Structural analysis of an epsilon-class glutathione transferase from housefly, Musca domestica.

Glutathione transferases (GSTs) play an important role in the detoxification of insecticides, and as such, they are a key contributor to enhanced resistance to insecticides. In the housefly (Musca domestica), two epsilon-class GSTs (MdGST6A and MdGST6B) that share high sequence homology have been identified, which are believed to be involved in resistance against insecticides. The structural determinants controlling the substrate specificity and enzyme activity of MdGST6s are unknown. The aim of this study was to crystallize and perform structural analysis of the GST isozyme, MdGST6B. The crystal structure of MdGST6B complexed with reduced glutathione (GSH) was determined at a resolution of 1.8 Å. MdGST6B was found to have a typical GST folding comprised of N-terminal and C-terminal domains. Arg113 and Phe121 on helix 4 were shown to protrude into the substrate binding pocket, and as a result, the entrance of the substrate binding pocket was narrower compared to delta- and epsilon-class GSTs from Africa malaria vector Anopheles gambiae, agGSTd1-6 and agGSTe2, respectively. This substrate pocket narrowing is partly due to the presence of a π-helix in the middle of helix 4. Among the six residues that donate hydrogen bonds to GSH, only Arg113 was located in the C-terminal domain. Ala substitution of Arg113 did not have a significant effect on enzyme activity, suggesting that the Arg113 hydrogen bond does not play a crucial role in catalysis. On the other hand, mutation at Phe108, located just below Arg113 in the binding pocket, reduced the affinity and catalytic activity to both GSH and the electrophilic co-substrate, 1-chloro-2,4-dinitrobenzene.

Design, synthesis and biological evaluation of organophosphorous-homodimers as dual binding site acetylcholinesterase inhibitors.

The cluster effect is an effective strategy to explore new lead compounds, and has been successfully applied in rational drug design and screening. A series of novel organophosphorous-homodimers were designed and synthesized based on the dual-site structure characteristics of acetylcholinesterase (AChE). The compounds were evaluated in vitro for their inhibitory activity to AChE extracted from Drosophila melanogaster and Musca domestic. Compound 4H showed an excellent inhibitor activity to both Drosophila melanogaster and Musca domestic with the corresponding IC(50) values of 23 and 168 nM, respectively. Meanwhile, its activities against Drosophila melanogaster and Musca domestic AChE were more than 10,00,000 and 100,000-fold higher compared with the parent compound (MH), and was up to 245 and 107-fold higher than those of the positive control omethoate. The molecular docking study revealed that 4H possessed an optimal spacer length and can perfectly fit into the central pocket, active gorge, and peripheral site of DmAChE, and consequently exhibited highly improved inhibitor potency to DmAChE. The bioassay tests showed that 4 series compounds showed prominent insecticidal activities against both Lipaphser erysimi and Tetranychus cinnbarinus at a concentration of 200mg/L. The insecticide activity of compound 4H was particularly significant that can cause 96% mortality to Tetranychus cinnbarinus after 24h of treatment.

[Tissue localization and expression difference of endogenous beta-glucosidase in digestive system of Musca domestica third instar larvae].

OBJECTIVE: To study the tissue localization and expression difference of endogenous beta-glucosidase in digestive system of Musca domestica third instar larvae. METHODS: The digestive system of the 3rd instar larvae of Musca domestic was taken for the below tests. Tissue localization of endogenous beta-glucosidase mRNA was identified by in situ hybridization. Cellulase was localized by immunohistochemistry. The enzymatic activity of beta-glucosidase was measured by 3, 5-dinitrosalicylic acid(DNS) assay. The relative mRNA expression levels of M. domestica beta-glucosidase gene in these organs were determined by RT-PCR. RESULTS: Beta-glucosidase mRNA, with in situ hybridization, was shown in the epithelial cells of midgut, salivary glands and foregut of the larvae. The immunohistochemical analysis on larvae tissues revealed that cellulase was produced and secreted by the epithelial cells of the midgut, salivary glands and foregut. beta-glucosidase activity in salivary glands, foregut, midgut, and hindgut was (0.80 +/- 0.06), (0.38 +/- 0.02), (1.20 +/- 0.05) and (0.26 +/- 0.02) IU/mg, respectively. There was significant difference in beta-glucosidase activity among these digestive organs (P < 0.05). The activity level of beta-glucosidase was highest in midgut [(45.45 +/- 1.27)%], and lowest in hindgut [(9.85 +/- 0.88)%]. However, beta-glucosidase gene were only expressed in the salivary gland, foregut and midgut. Significant differences in gene expression level of beta-glucosidase was found among these organs (P < 0.05). The relative expression quantity of beta-glucosidase gene in midgut and salivary glands were 5 and 3 times higher than that in foregut. CONCLUSION: The endogenous beta-glucosidase gene is expressed in the foregut, midgut and salivary glands. The midgut and salivary glands of Musca domestica 3rd instar larvae are the primary organs of this enzyme secretion.

Seasonal genetic variation in house fly populations, Musca domestica (Diptera: Muscidae).

Seasonal genetic variation was assessed in the common house fly Musca domestica. Allozymes at six gene enzyme system viz., ACPH, EST, G6PD, ME, AO and XDH were analyzed. Fourteen loci with twenty seven alleles were unraveled. The genetic variations were found to be affected to a great extent by environmental influence .F statistics has been used to calculate genetic variation which revealed that very little genetic variation has occurred among the house fly populations analyzed in the present study. Further, except ACPH-2, G6PD-2 and XDH-1 all the other loci show inbreeding (Fis>Fst). Thus it appears that the house fly populations analyzed are characterized by high level of inbreeding. Nei's genetic identity (I) and distance (D) values reveal a close similarity between summer and rainy season collections.

Cloning and molecular characterization of an ornithine decarboxylase gene and its expression during embryonic development of the housefly, Musca domestica.

We are interested in identifying targets that may be used to develop new control products for the common housefly, Musca domestica, a vector of disease for many vertebrates. One such target, ornithine decarboxylase (ODC), is an embryonic enzyme involved in the regulation of polyamines and is a critical enzyme during M. domestica development. In this study, the cDNA for ODC from M. domestica was cloned, sequenced, and characterized. The full-length cDNA was 1,337-bp, consistent with a single band of approximately 1.35 kb obtained by northern analysis. The open-reading frame contains 1,191 bp, yielding a deduced polypeptide of 396 amino acid residues with a predicted mass of 44,618 Da. The deduced M. domestica ODC protein was homologous to other ODC proteins. mRNA expression profiles analyzed by real-time PCR indicated that the ODC transcript is temporally regulated throughout embryogenesis. Sequence data and Southern blot analysis suggests that there were likely only one or two closely linked copies of the M. domestica ODC gene.

Enhanced pesticide sensitivity of novel housefly acetylcholinesterases: a new tool for the detection of residual pesticide contamination.

The full-length cDNA encoding an acetylcholinesterase (AChE) was cloned and sequenced from the housefly, Musca domestica, by reverse transcriptase-polymerase chain reaction (RT-PCR). Sequence analysis revealed that this 2,076 bp sequence encodes a mature protein of 612 amino acids (67 kDa) and a 79 residue signal peptide. The amino acid sequence shared 52.8-81.4% identity with the AChE proteins of other insects. The cDNA sequence, which lacked the signal peptide was inserted into the vector pPIC9K and then introduced into strain GS115 of the yeast Pichia pastoris. The recombinant AChE protein was then expressed in P. pastoris strain GS115 by methanol induction. Site-directed mutagenesis of the A262G, Y327F, Y327D and I374D residues, either singly or in combination, was performed by reverse PCR. These mutants improved the catalytic activity and sensitivity to the organophosphate and carbamate insecticides. Although the sensitivity of other mutants was slightly increased, the results still showed that the sensitivity of triple mutant, GDD (A262G/Y327D/I374D), enhanced remarkably as much as 16 times for methomyl, 14 times for both carbofuran and chlorpyrifos, and ten times for parathion-methyl, compared to that of the wild-type. The results strongly suggested that these residues are the key structural elements controlling AChE enzyme catalytic activity and sensitivity to inhibition by insecticides. The AChE enzyme obtained by this method could be used to detect the organophosphate and carbamate insecticide residues in fruits and vegetables, a characteristic of great potential research and industrial application.

PAP1 activates the prophenoloxidase system against bacterial infection in Musca domestica.

We previously identified three putative prophenoloxidase-activating proteinase (mdPAP1, mdPAP2, and mdPAP3) genes from housefly Musca domestica by transcriptomic analysis. In this study, mdPAP1 cDNA was cloned, and the function of its encoded protein was analyzed. The cDNA of mdPAP1 was 1358 bp, and it contained a single open reading frame of 1122 bp encoding a predicted MdPAP1 protein of 373 amino acids. The estimated molecular weight of MdPAP1 was 41267.08 Da with an isoelectric point of 6.25. The deduced amino acid sequence of MdPAP1 exhibited high similarity to known PAPs of insects. mdPAP1 was detected in larvae, pupae, and adult housefly, and the expression level of mdPAP1 was upregulated in bacterial challenged larvae. The recombinant protein of MdPAP1 expressed in Escherichia coli could cleave the prophenoloxidase into phenoloxidase in M. domestica hemolymph infected by bacteria and result in a significant increase of the total phenoloxidase activity. In addition, RNA interference-mediated gene silencing of mdPAP1 significantly increased the mortality of M. domestica larvae. Results indicated that mdPAP1 was involved in the activation of the prophenoloxidase against bacterial infection in M. domestica.

Expression, purification and direct eletrochemistry of cytochrome P450 6A1 from the house fly, Musca domestica.

A plasmid (pCW) was modified to code for the complete sequence of house fly (Musca domestica) cytochrome P450 6A1 (CYP6A1) with only the second amino acid changed in the N-terminal portion and this plasmid was used to express the enzyme CYP6A1 in Escherichia coli cells. With the addition of delta-aminolevulinic acid and FeCl(3) to the culture, the enzyme was produced at a level about 0.25 micromol L(-1) (15mgL(-1)) of culture with approximately 50% of the P450 being associated with the membrane fraction. The CYP6A1 protein was characterized and the content of CYP6A1 in each fraction was determined by the spectroscopic method. A nearly homogenous CYP6A1 was obtained by purification with a combination of DEAE Sepharose fast flow and hydroxyapatite chromatography. Direct electrochemistry of CYP6A1 in a didodecyldimethylammonium bromide (DSAB) film on an edge-plane pyrolytic graphite electrode (EPG) has been obtained and the catalytic activity of the enzyme to aldrin has been demonstrated by the cyclic voltammetry.

Activities of Select Enzymes Involved in Insecticide Resistance in Spinosad-Resistant and -Susceptible Strains of Musca domestica (Diptera: Muscidae).

A Musca domestica L. strain collected from Pakistan has recently been shown to be resistant to spinosad; however, there is scarce information about the mechanism of resistance. For this reason, we explored whether a metabolic-based mechanism was responsible by analyzing the activities of the metabolic detoxifying enzymes, carboxylesterases, glutathione S-transferases, and mixed-function oxidases, in both a spinosad-selected (Spin-SEL) strain of M. domestica and a susceptible counterpart (Lab-susceptible). The results revealed that both strains were statistically at par in terms of enzyme activities. The activity of carboxylesterases in the Lab-susceptible strain was 78.17 ± 3.06 in comparison to 79.16 ± 3.31 nmol min-1 mg-1 in the Spin-SEL strain. The activity of mixed-function oxidases was 51.58 ± 4.20 in the Lab-susceptible strain, whereas 54.33 ± 4.08 pmol min-1 mg-1 was recorded in the Spin-SEL strain. The activity of glutathione S-transferases was 86.50 ± 4.59 (Lab-susceptible) and 90.33 ± 2.81 nmol min-1 mg-1 (Spin-SEL). These results revealed that the studied enzymes might not be responsible for spinosad resistance in the studied strain of M. domestica. Therefore, studies should be extended to find out other possible mechanisms of spinosad resistance.

Ultra-highly sensitive organophosphorus biosensor based on chitosan/tin disulfide and British housefly acetylcholinesterase.

Pesticides have been extensively applied worldwide to protect crops from worms and insects; however, the continuous use of pesticides affects ecosystems, agricultural product safety, nontarget organisms, and human health. In this paper, we report a highly sensitive biosensor for the determination of pesticides based on tin sulfide (SnS2) and chitosan (CHIT) nanocomposites decorated with a unique British housefly acetylcholinesterase (AChE). The hydrothermally synthesized nano-SnS2 mixed with chitosan solution (CHIT-SnS2) was drop-casted onto a glassy carbon electrode (GCE). Subsequently, the British housefly AChE was immobilized on the CHIT/SnS2-coated GCE that was then employed for pesticide detection. The developed biosensor showed an ultra-high sensitivity and wide linear detection range from 0.02 nM to 20000 nM with a detection limit of 0.02 nM for the detection of chlorpyrifos as the model pesticide. Furthermore, the AChE/CHIT-SnS2/GCE exhibited acceptable storage stability, good reproducibility, and selectivity.

Reactivation potency of fluorinated pyridinium oximes for acetylcholinesterases inhibited by paraoxon organophosphorus agent.

For the purpose of developing new oxime reactivators of acetylcholinesterases (AChE) that have been inhibited by organophosphorus agents, emphasis was given to the finding that the lipophilic nature of fluorinated compounds is responsible for their enhanced transport across the blood brain barrier (BBB). As a result, we have designed and synthesized the fluorinated oxime derivatives, which quantum mechanical calculations suggest should have a greater lipophilicity and BBB permeability than their non-fluorinated analogs. Among the compounds explored in this study, 4 was found to have the highest potency for reactivation of paraoxon-inhibited housefly (HF) AChE.

In silico approaches to evaluate the molecular properties of organophosphate compounds to inhibit acetylcholinesterase activity in housefly.

Organophosphate compounds (OPC) have become the primary choice as insecticides and are widely used across the world. Additionally, OPCs were also commonly used as a chemical warfare agent that triggers a great challenge to public safety. Exposure of OPCs to human causes immediate excitation of cholinergic neurotransmission through transient elevation of synaptic acetylcholine (ACh) levels and accumulations. Likewise, prolonged exposure of OPCs can affect the processes in immune response, carbohydrate metabolism, cardiovascular toxicity, and several others. Studies revealed that the toxicity of OPCs was provoked by inhibition of acetylcholinesterase (AChE). Therefore, combined in silico approaches - pharmacophore-based 3D-QSAR model; docking and Molecular Dynamics (MD) - were used to assess the precise and comprehensive effects of series of known OP-derived compounds together with its -log LD50 values. The selected five-featured pharmacophore model - AAHHR.61 - displayed the highest correlation (R2 = .9166), cross-validated coefficient (Q2 = .8221), F = 63.2, Pearson-R = .9615 with low RMSE = .2621 values obtained using five component PLS factors. Subsequently, the well-validated model was then used as a 3D query to search novel OPCs using a high-throughput virtual screening technique. Simultaneously, the docking studies predicted the binding pose of the most active OPC in the MdAChE binding pocket. Additionally, the stability of docking was verified using MD simulation. The results revealed that OP22 and predicted lead compounds bound tightly to S315 of MdAChE through potential hydrogen bond interaction over time. Overall, this study might provide valuable insight into binding mode of OPCs and hit compounds to inhibit AChE in housefly.

MdRDH1, a HSP67B2-like rhodanese homologue plays a positive role in maintaining redox balance in Musca domestica.

Rhodanese homology domains (RHODs) are the structural modules of ubiquitous tertiary that occur in three major evolutionary phyla. Despite the versatile and important physiological functions of RHODs containing proteins, little is known about their invertebrate counterparts. A novel HSP67B2-like single-domain rhodanese homologue, MdRDH1 from Musca domestica, whose expression can be induced by bacterial infection or oxidative stress. Silencing MdRDH1 through RNAi causes important accumulations of reactive oxygen species (ROS) and malondialdehyde (MDA), and increases mortality in the larvae treated with bacterial invasion. The E. coli with MdRDH1 and the mutant MdRDH1C135A are transformed, with significant rhodanese activity of the recombinant protein of MdRDH1 in vitro found, without no detection of enzyme activity of the mutant MdRDH1C135A, revealing that catalytic Cys135 in the active-site loop is essential in the sulfurtransferase activity of MdRDH1. When oxidative stress is insulted by phenazine methosulfate (PMS), the MdRDH1 transformed E. coli shows enhanced survival rates compared with those bacteria transformed with MdRDH1C135A. Our research indicates that MdRDH1 confers oxidative stress tolerance, thus rendering evidence for the idea that rhodanese family genes play a critical role in antioxidant defenses. This paper yields novel insights into the potential antioxidative and immune functions of HSP67B2-like rhodanese homologues in invertebrate.