Cloning, recombinant expression, and characterization of a Rhipicephalus microplus carboxylesterase. [version 1; peer review: 2 approved, 1 not approved]
Background The Rhipicephalus microplus carboxylesterase (CBE) is involved in synthetic pyrethroid (SP) hydrolysis and historic evidence suggests that a non-synonymous mutation (Asp374Asn) in CBE is associated with increased resistance towards SP-based acaricides. Functional expression and characteri...
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| Language: | English |
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F1000 Research Ltd
2024-08-01
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| Series: | Gates Open Research |
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| Online Access: | https://gatesopenresearch.org/articles/8-87/v1 |
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| _version_ | 1841555781675122688 |
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| author | Michel Labuschagne |
| author_facet | Michel Labuschagne |
| author_sort | Michel Labuschagne |
| collection | DOAJ |
| description | Background The Rhipicephalus microplus carboxylesterase (CBE) is involved in synthetic pyrethroid (SP) hydrolysis and historic evidence suggests that a non-synonymous mutation (Asp374Asn) in CBE is associated with increased resistance towards SP-based acaricides. Functional expression and characterization of the wild-type and mutant CBE is required to understand the impact of the mutation on SP-based resistance. Methods The R. microplus CBE gene was cloned and functionally expressed in Pichia pastoris following the removal of the native signal peptide. Site directed mutagenesis was used to introduce the Asp374Asn substitution. Results Functional expression, characterization, and purification of both wild-type and mutant R. microplus CBE proteins was achieved using affinity chromatography under native conditions. Conclusions This report provides the necessary information for the tick research community to produce recombinant tick derived CBE proteins and to characterize the recombinant proteins towards substrates of interest. |
| format | Article |
| id | doaj-art-cde0cfb158d843e28b733febafa922d0 |
| institution | Kabale University |
| issn | 2572-4754 |
| language | English |
| publishDate | 2024-08-01 |
| publisher | F1000 Research Ltd |
| record_format | Article |
| series | Gates Open Research |
| spelling | doaj-art-cde0cfb158d843e28b733febafa922d02025-01-08T01:00:00ZengF1000 Research LtdGates Open Research2572-47542024-08-01817609Cloning, recombinant expression, and characterization of a Rhipicephalus microplus carboxylesterase. [version 1; peer review: 2 approved, 1 not approved]Michel Labuschagne0https://orcid.org/0000-0001-7758-3855Research Innovation, Clinglobal, Tamarin, 90903, MauritiusBackground The Rhipicephalus microplus carboxylesterase (CBE) is involved in synthetic pyrethroid (SP) hydrolysis and historic evidence suggests that a non-synonymous mutation (Asp374Asn) in CBE is associated with increased resistance towards SP-based acaricides. Functional expression and characterization of the wild-type and mutant CBE is required to understand the impact of the mutation on SP-based resistance. Methods The R. microplus CBE gene was cloned and functionally expressed in Pichia pastoris following the removal of the native signal peptide. Site directed mutagenesis was used to introduce the Asp374Asn substitution. Results Functional expression, characterization, and purification of both wild-type and mutant R. microplus CBE proteins was achieved using affinity chromatography under native conditions. Conclusions This report provides the necessary information for the tick research community to produce recombinant tick derived CBE proteins and to characterize the recombinant proteins towards substrates of interest.https://gatesopenresearch.org/articles/8-87/v1Rhipicephalus microplus; Carboxylesterase; Recombinant expression; Purification; Resistance; Pichia pastoriseng |
| spellingShingle | Michel Labuschagne Cloning, recombinant expression, and characterization of a Rhipicephalus microplus carboxylesterase. [version 1; peer review: 2 approved, 1 not approved] Gates Open Research Rhipicephalus microplus; Carboxylesterase; Recombinant expression; Purification; Resistance; Pichia pastoris eng |
| title | Cloning, recombinant expression, and characterization of a Rhipicephalus microplus carboxylesterase. [version 1; peer review: 2 approved, 1 not approved] |
| title_full | Cloning, recombinant expression, and characterization of a Rhipicephalus microplus carboxylesterase. [version 1; peer review: 2 approved, 1 not approved] |
| title_fullStr | Cloning, recombinant expression, and characterization of a Rhipicephalus microplus carboxylesterase. [version 1; peer review: 2 approved, 1 not approved] |
| title_full_unstemmed | Cloning, recombinant expression, and characterization of a Rhipicephalus microplus carboxylesterase. [version 1; peer review: 2 approved, 1 not approved] |
| title_short | Cloning, recombinant expression, and characterization of a Rhipicephalus microplus carboxylesterase. [version 1; peer review: 2 approved, 1 not approved] |
| title_sort | cloning recombinant expression and characterization of a rhipicephalus microplus carboxylesterase version 1 peer review 2 approved 1 not approved |
| topic | Rhipicephalus microplus; Carboxylesterase; Recombinant expression; Purification; Resistance; Pichia pastoris eng |
| url | https://gatesopenresearch.org/articles/8-87/v1 |
| work_keys_str_mv | AT michellabuschagne cloningrecombinantexpressionandcharacterizationofarhipicephalusmicropluscarboxylesteraseversion1peerreview2approved1notapproved |