Synergistic codon optimization and bioreactor cultivation toward enhanced secretion of fungal lignin peroxidase in Pichia pastoris: Enzymatic valorization of technical (industrial) lignins.
Identifieur interne : 000009 ( Main/Exploration ); précédent : 000008; suivant : 000010Synergistic codon optimization and bioreactor cultivation toward enhanced secretion of fungal lignin peroxidase in Pichia pastoris: Enzymatic valorization of technical (industrial) lignins.
Auteurs : B M Majeke [Afrique du Sud] ; M. García-Aparicio [Afrique du Sud] ; O D Biko [Afrique du Sud] ; M. Viljoen-Bloom [Afrique du Sud] ; W H Van Zyl [Afrique du Sud] ; J F Görgens [Afrique du Sud]Source :
- Enzyme and microbial technology [ 1879-0909 ] ; 2020.
Abstract
Lignin peroxidase (LiP) is a well-recognized enzyme for its ability to oxidize lignins, but its commercial availability is limited, which hinders the biotechnological application of LiP-based bioprocesses in lignocellulose biorefineries. This study evaluated a combination strategy to improve the expression of LiP to promote its practical use. The strategy included optimization of the lipH8 gene of Phanerochaete chrysosporium according to the codon usage of Pichia pastoris, followed by fed-batch fermentation using a 14 L bioreactor (10 L working volume). The combination strategy achieved a maximum volumetric LiPH8 activity of 4480 U L-1, protein concentration of 417 mg L-1 and a specific activity of 10.7 U mg-1, which was higher than previous reports. Biochemical characterization showed that the recombinant LiPH8 (rLiPH8) was optimum at pH 3.0, 25 ℃ and 0.4 mM H2O2. Using the optimized conditions, rLiPH8 was used to treat isolated technical lignins namely soda-anthraquinone (SAQ) lignin and steam explosion (S-E) lignin. High-performance gel permeation chromatography (HP-GPC) analysis showed that the molecular weight (Mw) of SAQ and S-E lignins were increased by 1.43-and 1.14-fold, respectively, after the enzymatic treatment. Thermogravimetric analysis (TGA) also showed that the thermal stability of the lignins was improved, indicating that the enzyme treatment of lignins with rLiPH8 resulted in lignin re-polymerization. As the first report on rLiPH8 production using P. pastoris, this study has shed light on the possible route for the enhancement of rLiPH8 production and its potential application for upgrading technical lignins.
DOI: 10.1016/j.enzmictec.2020.109593
PubMed: 32732041
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Electronic address: 16628233@sun.ac.za.</nlm:affiliation><country xml:lang="fr">Afrique du Sud</country><wicri:regionArea>Department of Process Engineering, Stellenbosch University, Private Bag X1, Matieland 7602</wicri:regionArea><wicri:noRegion>Matieland 7602</wicri:noRegion></affiliation></author><author><name sortKey="Garcia Aparicio, M" sort="Garcia Aparicio, M" uniqKey="Garcia Aparicio M" first="M" last="García-Aparicio">M. García-Aparicio</name><affiliation wicri:level="1"><nlm:affiliation>Department of Process Engineering, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa; Department of Microbiology, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. Electronic address: garcia@sun.ac.za.</nlm:affiliation><country xml:lang="fr">Afrique du Sud</country><wicri:regionArea>Department of Process Engineering, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa; Department of Microbiology, Stellenbosch University, Private Bag X1, Matieland 7602</wicri:regionArea><wicri:noRegion>Matieland 7602</wicri:noRegion></affiliation></author><author><name sortKey="Biko, O D" sort="Biko, O D" uniqKey="Biko O" first="O D" last="Biko">O D Biko</name><affiliation wicri:level="1"><nlm:affiliation>Department of Microbiology, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. Electronic address: 20657447@sun.ac.za.</nlm:affiliation><country xml:lang="fr">Afrique du Sud</country><wicri:regionArea>Department of Microbiology, Stellenbosch University, Private Bag X1, Matieland 7602</wicri:regionArea><wicri:noRegion>Matieland 7602</wicri:noRegion></affiliation></author><author><name sortKey="Viljoen Bloom, M" sort="Viljoen Bloom, M" uniqKey="Viljoen Bloom M" first="M" last="Viljoen-Bloom">M. Viljoen-Bloom</name><affiliation wicri:level="1"><nlm:affiliation>Department of Microbiology, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. 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Electronic address: jgorgens@sun.ac.za.</nlm:affiliation><country xml:lang="fr">Afrique du Sud</country><wicri:regionArea>Department of Process Engineering, Stellenbosch University, Private Bag X1, Matieland 7602</wicri:regionArea><wicri:noRegion>Matieland 7602</wicri:noRegion></affiliation></author></analytic><series><title level="j">Enzyme and microbial technology</title><idno type="eISSN">1879-0909</idno><imprint><date when="2020" type="published">2020</date></imprint></series></biblStruct></sourceDesc></fileDesc><profileDesc><textClass></textClass></profileDesc></teiHeader><front><div type="abstract" xml:lang="en">Lignin peroxidase (LiP) is a well-recognized enzyme for its ability to oxidize lignins, but its commercial availability is limited, which hinders the biotechnological application of LiP-based bioprocesses in lignocellulose biorefineries. This study evaluated a combination strategy to improve the expression of LiP to promote its practical use. The strategy included optimization of the lipH8 gene of Phanerochaete chrysosporium according to the codon usage of Pichia pastoris, followed by fed-batch fermentation using a 14 L bioreactor (10 L working volume). The combination strategy achieved a maximum volumetric LiPH8 activity of 4480 U L<sup>-1</sup>, protein concentration of 417 mg L<sup>-1</sup> and a specific activity of 10.7 U mg<sup>-1</sup>, which was higher than previous reports. Biochemical characterization showed that the recombinant LiPH8 (rLiPH8) was optimum at pH 3.0, 25 ℃ and 0.4 mM H<sub>2</sub>O<sub>2</sub>. Using the optimized conditions, rLiPH8 was used to treat isolated technical lignins namely soda-anthraquinone (SAQ) lignin and steam explosion (S-E) lignin. High-performance gel permeation chromatography (HP-GPC) analysis showed that the molecular weight (Mw) of SAQ and S-E lignins were increased by 1.43-and 1.14-fold, respectively, after the enzymatic treatment. Thermogravimetric analysis (TGA) also showed that the thermal stability of the lignins was improved, indicating that the enzyme treatment of lignins with rLiPH8 resulted in lignin re-polymerization. As the first report on rLiPH8 production using P. pastoris, this study has shed light on the possible route for the enhancement of rLiPH8 production and its potential application for upgrading technical lignins.</div></front></TEI><pubmed><MedlineCitation Status="In-Process" Owner="NLM"><PMID Version="1">32732041</PMID><DateRevised><Year>2020</Year><Month>07</Month><Day>31</Day></DateRevised><Article PubModel="Print-Electronic"><Journal><ISSN IssnType="Electronic">1879-0909</ISSN><JournalIssue CitedMedium="Internet"><Volume>139</Volume><PubDate><Year>2020</Year><Month>Sep</Month></PubDate></JournalIssue><Title>Enzyme and microbial technology</Title><ISOAbbreviation>Enzyme Microb Technol</ISOAbbreviation></Journal><ArticleTitle>Synergistic codon optimization and bioreactor cultivation toward enhanced secretion of fungal lignin peroxidase in Pichia pastoris: Enzymatic valorization of technical (industrial) lignins.</ArticleTitle><Pagination><MedlinePgn>109593</MedlinePgn></Pagination><ELocationID EIdType="pii" ValidYN="Y">S0141-0229(20)30086-7</ELocationID><ELocationID EIdType="doi" ValidYN="Y">10.1016/j.enzmictec.2020.109593</ELocationID><Abstract><AbstractText>Lignin peroxidase (LiP) is a well-recognized enzyme for its ability to oxidize lignins, but its commercial availability is limited, which hinders the biotechnological application of LiP-based bioprocesses in lignocellulose biorefineries. This study evaluated a combination strategy to improve the expression of LiP to promote its practical use. The strategy included optimization of the lipH8 gene of Phanerochaete chrysosporium according to the codon usage of Pichia pastoris, followed by fed-batch fermentation using a 14 L bioreactor (10 L working volume). The combination strategy achieved a maximum volumetric LiPH8 activity of 4480 U L<sup>-1</sup>, protein concentration of 417 mg L<sup>-1</sup> and a specific activity of 10.7 U mg<sup>-1</sup>, which was higher than previous reports. Biochemical characterization showed that the recombinant LiPH8 (rLiPH8) was optimum at pH 3.0, 25 ℃ and 0.4 mM H<sub>2</sub>O<sub>2</sub>. Using the optimized conditions, rLiPH8 was used to treat isolated technical lignins namely soda-anthraquinone (SAQ) lignin and steam explosion (S-E) lignin. High-performance gel permeation chromatography (HP-GPC) analysis showed that the molecular weight (Mw) of SAQ and S-E lignins were increased by 1.43-and 1.14-fold, respectively, after the enzymatic treatment. Thermogravimetric analysis (TGA) also showed that the thermal stability of the lignins was improved, indicating that the enzyme treatment of lignins with rLiPH8 resulted in lignin re-polymerization. As the first report on rLiPH8 production using P. pastoris, this study has shed light on the possible route for the enhancement of rLiPH8 production and its potential application for upgrading technical lignins.</AbstractText><CopyrightInformation>Copyright © 2020 Elsevier Inc. All rights reserved.</CopyrightInformation></Abstract><AuthorList CompleteYN="Y"><Author ValidYN="Y"><LastName>Majeke</LastName><ForeName>B M</ForeName><Initials>BM</Initials><AffiliationInfo><Affiliation>Department of Process Engineering, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. Electronic address: 16628233@sun.ac.za.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>García-Aparicio</LastName><ForeName>M</ForeName><Initials>M</Initials><AffiliationInfo><Affiliation>Department of Process Engineering, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa; Department of Microbiology, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. Electronic address: garcia@sun.ac.za.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Biko</LastName><ForeName>O D</ForeName><Initials>OD</Initials><AffiliationInfo><Affiliation>Department of Microbiology, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. Electronic address: 20657447@sun.ac.za.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Viljoen-Bloom</LastName><ForeName>M</ForeName><Initials>M</Initials><AffiliationInfo><Affiliation>Department of Microbiology, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. Electronic address: mv4@sun.ac.za.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>van Zyl</LastName><ForeName>W H</ForeName><Initials>WH</Initials><AffiliationInfo><Affiliation>Department of Microbiology, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. Electronic address: whvz@sun.ac.za.</Affiliation></AffiliationInfo></Author><Author ValidYN="Y"><LastName>Görgens</LastName><ForeName>J F</ForeName><Initials>JF</Initials><AffiliationInfo><Affiliation>Department of Process Engineering, Stellenbosch University, Private Bag X1, Matieland 7602, South Africa. Electronic address: jgorgens@sun.ac.za.</Affiliation></AffiliationInfo></Author></AuthorList><Language>eng</Language><PublicationTypeList><PublicationType UI="D016428">Journal Article</PublicationType></PublicationTypeList><ArticleDate DateType="Electronic"><Year>2020</Year><Month>05</Month><Day>12</Day></ArticleDate></Article><MedlineJournalInfo><Country>United States</Country><MedlineTA>Enzyme Microb Technol</MedlineTA><NlmUniqueID>8003761</NlmUniqueID><ISSNLinking>0141-0229</ISSNLinking></MedlineJournalInfo><CitationSubset>IM</CitationSubset><KeywordList Owner="NOTNLM"><Keyword MajorTopicYN="N">Codon optimization</Keyword><Keyword MajorTopicYN="N">Fed-batch fermentation</Keyword><Keyword MajorTopicYN="N">Lignin peroxidase</Keyword><Keyword MajorTopicYN="N">Lignin polymerization</Keyword><Keyword MajorTopicYN="N">Pichia pastoris</Keyword><Keyword MajorTopicYN="N">Technical lignin</Keyword></KeywordList><CoiStatement>Declaration of Competing Interest None.</CoiStatement></MedlineCitation><PubmedData><History><PubMedPubDate PubStatus="received"><Year>2020</Year><Month>03</Month><Day>18</Day></PubMedPubDate><PubMedPubDate PubStatus="revised"><Year>2020</Year><Month>05</Month><Day>01</Day></PubMedPubDate><PubMedPubDate PubStatus="accepted"><Year>2020</Year><Month>05</Month><Day>08</Day></PubMedPubDate><PubMedPubDate PubStatus="entrez"><Year>2020</Year><Month>8</Month><Day>1</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="pubmed"><Year>2020</Year><Month>8</Month><Day>1</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate><PubMedPubDate PubStatus="medline"><Year>2020</Year><Month>8</Month><Day>1</Day><Hour>6</Hour><Minute>0</Minute></PubMedPubDate></History><PublicationStatus>ppublish</PublicationStatus><ArticleIdList><ArticleId IdType="pubmed">32732041</ArticleId><ArticleId IdType="pii">S0141-0229(20)30086-7</ArticleId><ArticleId IdType="doi">10.1016/j.enzmictec.2020.109593</ArticleId></ArticleIdList></PubmedData></pubmed><affiliations><list><country><li>Afrique du Sud</li></country></list><tree><country name="Afrique du Sud"><noRegion><name sortKey="Majeke, B M" sort="Majeke, B M" uniqKey="Majeke B" first="B M" last="Majeke">B M Majeke</name></noRegion><name sortKey="Biko, O D" sort="Biko, O D" uniqKey="Biko O" first="O D" last="Biko">O D Biko</name><name sortKey="Garcia Aparicio, M" sort="Garcia Aparicio, M" uniqKey="Garcia Aparicio M" first="M" last="García-Aparicio">M. García-Aparicio</name><name sortKey="Gorgens, J F" sort="Gorgens, J F" uniqKey="Gorgens J" first="J F" last="Görgens">J F Görgens</name><name sortKey="Van Zyl, W H" sort="Van Zyl, W H" uniqKey="Van Zyl W" first="W H" last="Van Zyl">W H Van Zyl</name><name sortKey="Viljoen Bloom, M" sort="Viljoen Bloom, M" uniqKey="Viljoen Bloom M" first="M" last="Viljoen-Bloom">M. Viljoen-Bloom</name></country></tree></affiliations></record>Pour manipuler ce document sous Unix (Dilib)
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