Biochemical Characteristics and Variable Alginate-Degrading Modes of a Novel Bifunctional Endolytic Alginate Lyase
Bifunctional alginate lyases can efficiently degrade alginate comprised of mannuronate (M) and guluronate (G), but their substrate-degrading modes have not been thoroughly elucidated to date. In this study, we present Aly1 as a novel bifunctional endolytic alginate lyase of the genus The recombinant...
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description | Bifunctional alginate lyases can efficiently degrade alginate comprised of mannuronate (M) and guluronate (G), but their substrate-degrading modes have not been thoroughly elucidated to date. In this study, we present Aly1 as a novel bifunctional endolytic alginate lyase of the genus
The recombinant enzyme showed optimal activity at 50°C and pH 6.0. The enzyme produced unsaturated disaccharide (UDP2) and trisaccharide fractions as the final main alginate digests. Primary substrate preference tests and further structure identification of various size-defined final oligosaccharide products demonstrated that Aly1 is a bifunctional alginate lyase and prefers G to M. Tetrasaccharide-size fractions are the smallest substrates, and M, G, and UDP2 fractions are the minimal product types. Remarkably, Aly1 can vary its substrate-degrading modes in accordance with the terminus types, molecular sizes, and M/G contents of alginate substrates, producing a series of small size-defined saturated oligosaccharide products from the nonreducing ends of single or different saturated sugar chains and yielding unsaturated products in distinct but restricted patterns. The action mode changes can be partially inhibited by fluorescent labeling at the reducing ends of oligosaccharide substrates. Deletion of the noncatalytic region (NCR) of Aly1 caused weak changes of biochemical characteristics but increased the degradation proportions of small size-defined saturated M-enriched oligosaccharide substrates and unsaturated tetrasaccharide fractions without any size changes of degradable oligosaccharides, thereby enhancing the M preference and enzyme activity. Therefore, our results provided insight into the variable action mode of a novel bifunctional endolytic alginate lyase to inform accurate enzyme use.
The elucidated endolytic alginate lyases usually degrade substrates into various size-defined unsaturated oligosaccharide products (≥UDP2), and exolytic enzymes yield primarily unsaturated monosaccharide products. However, it is poorly understood whether endolytic enzymes can produce monosaccharide product types when degrading alginate. In this study, we demonstrated that Aly1, a bifunctional alginate lyase of
sp. strain MY04, is endolytic and monosaccharide producing. Using various sugar chains as testing substrates, we also proved that key factors causing Aly1's action mode changes are the terminus types, molecular sizes, and M/G contents of substrates. Furthermore, the NCR fragmen |
doi_str_mv | 10.1128/AEM.01608-17 |
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The recombinant enzyme showed optimal activity at 50°C and pH 6.0. The enzyme produced unsaturated disaccharide (UDP2) and trisaccharide fractions as the final main alginate digests. Primary substrate preference tests and further structure identification of various size-defined final oligosaccharide products demonstrated that Aly1 is a bifunctional alginate lyase and prefers G to M. Tetrasaccharide-size fractions are the smallest substrates, and M, G, and UDP2 fractions are the minimal product types. Remarkably, Aly1 can vary its substrate-degrading modes in accordance with the terminus types, molecular sizes, and M/G contents of alginate substrates, producing a series of small size-defined saturated oligosaccharide products from the nonreducing ends of single or different saturated sugar chains and yielding unsaturated products in distinct but restricted patterns. The action mode changes can be partially inhibited by fluorescent labeling at the reducing ends of oligosaccharide substrates. Deletion of the noncatalytic region (NCR) of Aly1 caused weak changes of biochemical characteristics but increased the degradation proportions of small size-defined saturated M-enriched oligosaccharide substrates and unsaturated tetrasaccharide fractions without any size changes of degradable oligosaccharides, thereby enhancing the M preference and enzyme activity. Therefore, our results provided insight into the variable action mode of a novel bifunctional endolytic alginate lyase to inform accurate enzyme use.
The elucidated endolytic alginate lyases usually degrade substrates into various size-defined unsaturated oligosaccharide products (≥UDP2), and exolytic enzymes yield primarily unsaturated monosaccharide products. However, it is poorly understood whether endolytic enzymes can produce monosaccharide product types when degrading alginate. In this study, we demonstrated that Aly1, a bifunctional alginate lyase of
sp. strain MY04, is endolytic and monosaccharide producing. Using various sugar chains as testing substrates, we also proved that key factors causing Aly1's action mode changes are the terminus types, molecular sizes, and M/G contents of substrates. Furthermore, the NCR fragment's effects on Aly1's biochemical characteristics and alginate-degrading modes and corresponding mechanisms were discovered by gene truncation and enzyme comparison. In summary, this study provides a novel bifunctional endolytic tool and a variable action mode for accurate use in alginate degradation.</description><identifier>ISSN: 0099-2240</identifier><identifier>EISSN: 1098-5336</identifier><identifier>DOI: 10.1128/AEM.01608-17</identifier><identifier>PMID: 28939598</identifier><language>eng</language><publisher>United States: American Society for Microbiology</publisher><subject>Alginate lyase ; Alginic acid ; Biochemical characteristics ; Biochemistry ; Degradation ; Enzymatic activity ; Enzyme activity ; Enzymes ; Enzymology and Protein Engineering ; Fluorescence ; Oligosaccharides ; pH effects ; Substrates ; Sugar</subject><ispartof>Applied and environmental microbiology, 2017-12, Vol.83 (23), p.1</ispartof><rights>Copyright © 2017 Cheng et al.</rights><rights>Copyright American Society for Microbiology Dec 2017</rights><rights>Copyright © 2017 Cheng et al. 2017 Cheng et al.</rights><lds50>peer_reviewed</lds50><oa>free_for_read</oa><woscitedreferencessubscribed>false</woscitedreferencessubscribed><citedby>FETCH-LOGICAL-c412t-46df4dd11cbf556cff309088547eadcfd677c400dd01a67bd4c5512500a744193</citedby><cites>FETCH-LOGICAL-c412t-46df4dd11cbf556cff309088547eadcfd677c400dd01a67bd4c5512500a744193</cites></display><links><openurl>$$Topenurl_article</openurl><openurlfulltext>$$Topenurlfull_article</openurlfulltext><thumbnail>$$Tsyndetics_thumb_exl</thumbnail><linktopdf>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5691422/pdf/$$EPDF$$P50$$Gpubmedcentral$$Hfree_for_read</linktopdf><linktohtml>$$Uhttps://www.ncbi.nlm.nih.gov/pmc/articles/PMC5691422/$$EHTML$$P50$$Gpubmedcentral$$Hfree_for_read</linktohtml><link.rule.ids>230,314,727,780,784,885,3188,27924,27925,53791,53793</link.rule.ids><backlink>$$Uhttps://www.ncbi.nlm.nih.gov/pubmed/28939598$$D View this record in MEDLINE/PubMed$$Hfree_for_read</backlink></links><search><contributor>Müller, Volker</contributor><creatorcontrib>Cheng, Yuanyuan</creatorcontrib><creatorcontrib>Wang, Dandan</creatorcontrib><creatorcontrib>Gu, Jingyan</creatorcontrib><creatorcontrib>Li, Junge</creatorcontrib><creatorcontrib>Liu, Huihui</creatorcontrib><creatorcontrib>Li, Fuchuan</creatorcontrib><creatorcontrib>Han, Wenjun</creatorcontrib><title>Biochemical Characteristics and Variable Alginate-Degrading Modes of a Novel Bifunctional Endolytic Alginate Lyase</title><title>Applied and environmental microbiology</title><addtitle>Appl Environ Microbiol</addtitle><description>Bifunctional alginate lyases can efficiently degrade alginate comprised of mannuronate (M) and guluronate (G), but their substrate-degrading modes have not been thoroughly elucidated to date. In this study, we present Aly1 as a novel bifunctional endolytic alginate lyase of the genus
The recombinant enzyme showed optimal activity at 50°C and pH 6.0. The enzyme produced unsaturated disaccharide (UDP2) and trisaccharide fractions as the final main alginate digests. Primary substrate preference tests and further structure identification of various size-defined final oligosaccharide products demonstrated that Aly1 is a bifunctional alginate lyase and prefers G to M. Tetrasaccharide-size fractions are the smallest substrates, and M, G, and UDP2 fractions are the minimal product types. Remarkably, Aly1 can vary its substrate-degrading modes in accordance with the terminus types, molecular sizes, and M/G contents of alginate substrates, producing a series of small size-defined saturated oligosaccharide products from the nonreducing ends of single or different saturated sugar chains and yielding unsaturated products in distinct but restricted patterns. The action mode changes can be partially inhibited by fluorescent labeling at the reducing ends of oligosaccharide substrates. Deletion of the noncatalytic region (NCR) of Aly1 caused weak changes of biochemical characteristics but increased the degradation proportions of small size-defined saturated M-enriched oligosaccharide substrates and unsaturated tetrasaccharide fractions without any size changes of degradable oligosaccharides, thereby enhancing the M preference and enzyme activity. Therefore, our results provided insight into the variable action mode of a novel bifunctional endolytic alginate lyase to inform accurate enzyme use.
The elucidated endolytic alginate lyases usually degrade substrates into various size-defined unsaturated oligosaccharide products (≥UDP2), and exolytic enzymes yield primarily unsaturated monosaccharide products. However, it is poorly understood whether endolytic enzymes can produce monosaccharide product types when degrading alginate. In this study, we demonstrated that Aly1, a bifunctional alginate lyase of
sp. strain MY04, is endolytic and monosaccharide producing. Using various sugar chains as testing substrates, we also proved that key factors causing Aly1's action mode changes are the terminus types, molecular sizes, and M/G contents of substrates. Furthermore, the NCR fragment's effects on Aly1's biochemical characteristics and alginate-degrading modes and corresponding mechanisms were discovered by gene truncation and enzyme comparison. In summary, this study provides a novel bifunctional endolytic tool and a variable action mode for accurate use in alginate degradation.</description><subject>Alginate lyase</subject><subject>Alginic acid</subject><subject>Biochemical characteristics</subject><subject>Biochemistry</subject><subject>Degradation</subject><subject>Enzymatic activity</subject><subject>Enzyme activity</subject><subject>Enzymes</subject><subject>Enzymology and Protein Engineering</subject><subject>Fluorescence</subject><subject>Oligosaccharides</subject><subject>pH effects</subject><subject>Substrates</subject><subject>Sugar</subject><issn>0099-2240</issn><issn>1098-5336</issn><fulltext>true</fulltext><rsrctype>article</rsrctype><creationdate>2017</creationdate><recordtype>article</recordtype><recordid>eNpdkU1vEzEQhi0EoqFw44wsceHAtmOv7bUvSGkIFCktF-BqOf5IXDnr1t6tlH_fDS0RcJrDPPNoZl6E3hI4I4TK8_ny6gyIANmQ7hmaEVCy4W0rnqMZgFINpQxO0KtabwCAgZAv0QmVqlVcyRkqFzHbrd9FaxJebE0xdvAl1iHaik3v8C9Tolknj-dpE3sz-Oaz3xTjYr_BV9n5inPABl_ne5_wRQxjb4eY-8m27F1O-0l0HMWrvan-NXoRTKr-zVM9RT-_LH8sLpvV96_fFvNVYxmhQ8OEC8w5Quw6cC5sCC0okJKzzhtngxNdZxmAc0CM6NaOWc4J5QCmY4yo9hR9evTejuudd9b3QzFJ35a4M2Wvs4n6304ft3qT7zUXijBKJ8GHJ0HJd6Ovg97Fan1Kpvd5rJooRjtgtO0m9P1_6E0ey_SFAyVBKNUSMlEfHylbcq3Fh-MyBPQhTD2FqX-HqclB-u7vA47wn_TaB2S1mzE</recordid><startdate>20171201</startdate><enddate>20171201</enddate><creator>Cheng, Yuanyuan</creator><creator>Wang, Dandan</creator><creator>Gu, Jingyan</creator><creator>Li, Junge</creator><creator>Liu, Huihui</creator><creator>Li, Fuchuan</creator><creator>Han, Wenjun</creator><general>American Society for Microbiology</general><scope>NPM</scope><scope>AAYXX</scope><scope>CITATION</scope><scope>7QL</scope><scope>7QO</scope><scope>7SN</scope><scope>7SS</scope><scope>7ST</scope><scope>7T7</scope><scope>7TM</scope><scope>7U9</scope><scope>8FD</scope><scope>C1K</scope><scope>FR3</scope><scope>H94</scope><scope>M7N</scope><scope>P64</scope><scope>RC3</scope><scope>SOI</scope><scope>7X8</scope><scope>5PM</scope></search><sort><creationdate>20171201</creationdate><title>Biochemical Characteristics and Variable Alginate-Degrading Modes of a Novel Bifunctional Endolytic Alginate Lyase</title><author>Cheng, Yuanyuan ; Wang, Dandan ; Gu, Jingyan ; Li, Junge ; Liu, Huihui ; Li, Fuchuan ; Han, Wenjun</author></sort><facets><frbrtype>5</frbrtype><frbrgroupid>cdi_FETCH-LOGICAL-c412t-46df4dd11cbf556cff309088547eadcfd677c400dd01a67bd4c5512500a744193</frbrgroupid><rsrctype>articles</rsrctype><prefilter>articles</prefilter><language>eng</language><creationdate>2017</creationdate><topic>Alginate lyase</topic><topic>Alginic acid</topic><topic>Biochemical characteristics</topic><topic>Biochemistry</topic><topic>Degradation</topic><topic>Enzymatic activity</topic><topic>Enzyme activity</topic><topic>Enzymes</topic><topic>Enzymology and Protein Engineering</topic><topic>Fluorescence</topic><topic>Oligosaccharides</topic><topic>pH effects</topic><topic>Substrates</topic><topic>Sugar</topic><toplevel>peer_reviewed</toplevel><toplevel>online_resources</toplevel><creatorcontrib>Cheng, Yuanyuan</creatorcontrib><creatorcontrib>Wang, Dandan</creatorcontrib><creatorcontrib>Gu, Jingyan</creatorcontrib><creatorcontrib>Li, Junge</creatorcontrib><creatorcontrib>Liu, Huihui</creatorcontrib><creatorcontrib>Li, Fuchuan</creatorcontrib><creatorcontrib>Han, Wenjun</creatorcontrib><collection>PubMed</collection><collection>CrossRef</collection><collection>Bacteriology Abstracts (Microbiology B)</collection><collection>Biotechnology Research Abstracts</collection><collection>Ecology Abstracts</collection><collection>Entomology Abstracts (Full archive)</collection><collection>Environment Abstracts</collection><collection>Industrial and Applied Microbiology Abstracts (Microbiology A)</collection><collection>Nucleic Acids Abstracts</collection><collection>Virology and AIDS Abstracts</collection><collection>Technology Research Database</collection><collection>Environmental Sciences and Pollution Management</collection><collection>Engineering Research Database</collection><collection>AIDS and Cancer Research Abstracts</collection><collection>Algology Mycology and Protozoology Abstracts (Microbiology C)</collection><collection>Biotechnology and BioEngineering Abstracts</collection><collection>Genetics Abstracts</collection><collection>Environment Abstracts</collection><collection>MEDLINE - Academic</collection><collection>PubMed Central (Full Participant titles)</collection><jtitle>Applied and environmental microbiology</jtitle></facets><delivery><delcategory>Remote Search Resource</delcategory><fulltext>fulltext</fulltext></delivery><addata><au>Cheng, Yuanyuan</au><au>Wang, Dandan</au><au>Gu, Jingyan</au><au>Li, Junge</au><au>Liu, Huihui</au><au>Li, Fuchuan</au><au>Han, Wenjun</au><au>Müller, Volker</au><format>journal</format><genre>article</genre><ristype>JOUR</ristype><atitle>Biochemical Characteristics and Variable Alginate-Degrading Modes of a Novel Bifunctional Endolytic Alginate Lyase</atitle><jtitle>Applied and environmental microbiology</jtitle><addtitle>Appl Environ Microbiol</addtitle><date>2017-12-01</date><risdate>2017</risdate><volume>83</volume><issue>23</issue><spage>1</spage><pages>1-</pages><issn>0099-2240</issn><eissn>1098-5336</eissn><abstract>Bifunctional alginate lyases can efficiently degrade alginate comprised of mannuronate (M) and guluronate (G), but their substrate-degrading modes have not been thoroughly elucidated to date. In this study, we present Aly1 as a novel bifunctional endolytic alginate lyase of the genus
The recombinant enzyme showed optimal activity at 50°C and pH 6.0. The enzyme produced unsaturated disaccharide (UDP2) and trisaccharide fractions as the final main alginate digests. Primary substrate preference tests and further structure identification of various size-defined final oligosaccharide products demonstrated that Aly1 is a bifunctional alginate lyase and prefers G to M. Tetrasaccharide-size fractions are the smallest substrates, and M, G, and UDP2 fractions are the minimal product types. Remarkably, Aly1 can vary its substrate-degrading modes in accordance with the terminus types, molecular sizes, and M/G contents of alginate substrates, producing a series of small size-defined saturated oligosaccharide products from the nonreducing ends of single or different saturated sugar chains and yielding unsaturated products in distinct but restricted patterns. The action mode changes can be partially inhibited by fluorescent labeling at the reducing ends of oligosaccharide substrates. Deletion of the noncatalytic region (NCR) of Aly1 caused weak changes of biochemical characteristics but increased the degradation proportions of small size-defined saturated M-enriched oligosaccharide substrates and unsaturated tetrasaccharide fractions without any size changes of degradable oligosaccharides, thereby enhancing the M preference and enzyme activity. Therefore, our results provided insight into the variable action mode of a novel bifunctional endolytic alginate lyase to inform accurate enzyme use.
The elucidated endolytic alginate lyases usually degrade substrates into various size-defined unsaturated oligosaccharide products (≥UDP2), and exolytic enzymes yield primarily unsaturated monosaccharide products. However, it is poorly understood whether endolytic enzymes can produce monosaccharide product types when degrading alginate. In this study, we demonstrated that Aly1, a bifunctional alginate lyase of
sp. strain MY04, is endolytic and monosaccharide producing. Using various sugar chains as testing substrates, we also proved that key factors causing Aly1's action mode changes are the terminus types, molecular sizes, and M/G contents of substrates. Furthermore, the NCR fragment's effects on Aly1's biochemical characteristics and alginate-degrading modes and corresponding mechanisms were discovered by gene truncation and enzyme comparison. In summary, this study provides a novel bifunctional endolytic tool and a variable action mode for accurate use in alginate degradation.</abstract><cop>United States</cop><pub>American Society for Microbiology</pub><pmid>28939598</pmid><doi>10.1128/AEM.01608-17</doi><oa>free_for_read</oa></addata></record> |
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subjects | Alginate lyase Alginic acid Biochemical characteristics Biochemistry Degradation Enzymatic activity Enzyme activity Enzymes Enzymology and Protein Engineering Fluorescence Oligosaccharides pH effects Substrates Sugar |
title | Biochemical Characteristics and Variable Alginate-Degrading Modes of a Novel Bifunctional Endolytic Alginate Lyase |
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