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Generation of a library of carbohydrate-active enzymes for plant biomass deconstruction

2022-04-05Artigo científico Acesso aberto
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dc.contributor.authorCardoso, Vânia
dc.contributor.authorBras, Joana L. A.
dc.contributor.authorCosta, Ines F.
dc.contributor.authorFerreira, Luis M. A.
dc.contributor.authorGama, Luis
dc.contributor.authorVincentelli, Renaud
dc.contributor.authorHenrissat, Bernard
dc.contributor.authorFontes, Carlos M.G.A.
dc.date.accessioned2022-05-22T22:38:43Z
dc.date.available2022-05-22T22:38:43Z
dc.date.issued2022-04-05
dc.descriptionÁreas de pesquisa: Biochemistry & Molecular Biology ; Chemistrypt_PT
dc.description.abstractIn nature, the deconstruction of plant carbohydrates is carried out by carbohydrate-active enzymes (CAZymes). A high-throughput (HTP) strategy was used to isolate and clone 1476 genes obtained from a diverse library of recombinant CAZymes covering a variety of sequence-based families, enzyme classes, and source organisms. All genes were successfully isolated by either PCR (61%) or gene synthesis (GS) (39%) and were subsequently cloned into Escherichia coli expression vectors. Most proteins (79%) were obtained at a good yield during recombinant expression. A significantly lower number (p < 0.01) of proteins from eukaryotic (57.7%) and archaeal (53.3%) origin were soluble compared to bacteria (79.7%). Genes obtained by GS gave a significantly lower number (p = 0.04) of soluble proteins while the green fluorescent protein tag improved protein solubility (p = 0.05). Finally, a relationship between the amino acid composition and protein solubility was observed. Thus, a lower percentage of non-polar and higher percentage of negatively charged amino acids in a protein may be a good predictor for higher protein solubility in E. coli. The HTP approach presented here is a powerful tool for producing recombinant CAZymes that can be used for future studies of plant cell wall degradation. Successful production and expression of soluble recombinant proteins at a high rate opens new possibilities for the high-throughput production of targets from limitless sourcespt_PT
dc.description.versioninfo:eu-repo/semantics/publishedVersionpt_PT
dc.identifier.citationCardoso V, Brás JLA, Costa IF, Ferreira LMA, Gama LT, Vincentelli R, Henrissat B, Fontes CMGA. 2022. Generation of a library of carbohydrate-active enzymes for plant biomass deconstruction. Internal Journal of Molecular Sciences, 23(7):4024. DOI 10.3390/ ijms23074024pt_PT
dc.identifier.doi10.3390/ ijms23074024pt_PT
dc.identifier.eissn1422-0067
dc.identifier.urihttp://hdl.handle.net/10400.5/24345
dc.language.isoengpt_PT
dc.peerreviewedyespt_PT
dc.publisherMDPIpt_PT
dc.relationPortugal 2020 - 47033pt_PT
dc.relation.publisherversionhttps://www.mdpi.com/1422-0067/23/7/4024pt_PT
dc.rights.urihttp://creativecommons.org/licenses/by/4.0/pt_PT
dc.subjectCarbohydrate-active enzymes (CAZymes)pt_PT
dc.subjectPlant biomasspt_PT
dc.subjectGene synthesispt_PT
dc.subjectPCRpt_PT
dc.subjectHigh-throughput (HTP) cloningpt_PT
dc.subjectHTP expressionpt_PT
dc.titleGeneration of a library of carbohydrate-active enzymes for plant biomass deconstructionpt_PT
dc.typejournal article
dspace.entity.typePublication
oaire.citation.conferencePlaceBasel, Switzerlandpt_PT
oaire.citation.titleInternal Journal of Molecular Sciencespt_PT
oaire.citation.volume23(7):4024pt_PT
person.familyNameCardoso Lopes
person.familyNameGama
person.familyNameFontes
person.givenNameVânia Alexandra da Silva
person.givenNameLuis
person.givenNameCarlos
person.identifier.ciencia-id121B-B572-2236
person.identifier.ciencia-idC71D-140F-E876
person.identifier.ciencia-idC01A-FCB3-7F99
person.identifier.orcid0000-0001-8389-7543
person.identifier.orcid0000-0003-3894-3488
person.identifier.orcid0000-0002-1219-9753
person.identifier.ridN-4172-2018
person.identifier.scopus-author-id36027387300
person.identifier.scopus-author-id7005517465
rcaap.rightsopenAccesspt_PT
rcaap.typearticlept_PT
relation.isAuthorOfPublication36b1e43a-c5eb-4aae-91d9-20145f62532c
relation.isAuthorOfPublicatione6ff858c-9615-437f-98a7-b61b4362834c
relation.isAuthorOfPublicationefb2611f-c4fd-4954-b932-653ae9792f2c
relation.isAuthorOfPublication.latestForDiscovery36b1e43a-c5eb-4aae-91d9-20145f62532c

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