23
DOI: https://doi.org/10.1186/s43141-020-00103-0
English
eng
Vadala, Bhuvan Shankar
Contributor
<div id="abs0005" style="margin: 0px; padding: 0px; color: rgb(31, 31, 31); font-family: ElsevierGulliver, Georgia, "Times New Roman", Times, STIXGeneral, "Cambria Math", "Lucida Sans Unicode", "Microsoft Sans Serif", "Segoe UI Symbol", "Arial Unicode MS", serif, sans-serif; font-size: 16px;"><h3 class="u-h4 u-margin-m-top u-margin-xs-bottom" id="sect0010" style="margin-right: 0px; margin-left: 0px; padding: 0px; margin-top: 24px !important; margin-bottom: 8px !important; font-size: 20px !important; line-height: 30px !important;">Background</h3><div class="u-margin-s-bottom" id="sp0040" style="margin-top: 0px; margin-right: 0px; margin-left: 0px; padding: 0px; margin-bottom: 16px !important;"><span style="margin: 0px; padding: 0px;"><a href="https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/cellulase" title="Learn more about Cellulases from ScienceDirect's AI-generated Topic Pages" class="topic-link" style="margin: 0px; padding: 0px; background-color: rgba(0, 0, 0, 0); word-break: break-word; text-decoration-line: underline; text-decoration-thickness: 1px; text-decoration-color: rgb(31, 31, 31); color: rgb(31, 31, 31); text-underline-offset: 1px;">Cellulases</a> are well known for their various industrial applications. They are naturally produced by different species of bacteria and fungi. Fermentation process of cellulase producers has limitation due to the high substrate cost required for cellulase induction and challenges to maintain the suitable condition for the respective cellulase production. Recombinant cellulase production could be the potential solution to these problems. In the current study, we investigated recombinant cellulase expression in </span><em style="margin: 0px; padding: 0px;">Escherichia coli</em> using cellulase gene from <span style="margin: 0px; padding: 0px;"><span style="margin: 0px; padding: 0px;"><a href="https://www.sciencedirect.com/topics/immunology-and-microbiology/bacillus-subtilis" title="Learn more about Bacillus subtilis from ScienceDirect's AI-generated Topic Pages" class="topic-link" style="margin: 0px; padding: 0px; background-color: rgba(0, 0, 0, 0); word-break: break-word; text-decoration-line: underline; text-decoration-thickness: 1px; text-decoration-color: rgb(31, 31, 31); color: rgb(31, 31, 31); text-underline-offset: 1px;">Bacillus subtilis</a></span></span>.</div></div><div id="abs0010" style="margin: 0px; padding: 0px; color: rgb(31, 31, 31); font-family: ElsevierGulliver, Georgia, "Times New Roman", Times, STIXGeneral, "Cambria Math", "Lucida Sans Unicode", "Microsoft Sans Serif", "Segoe UI Symbol", "Arial Unicode MS", serif, sans-serif; font-size: 16px;"><h3 class="u-h4 u-margin-m-top u-margin-xs-bottom" id="sect0015" style="margin-right: 0px; margin-left: 0px; padding: 0px; margin-top: 24px !important; margin-bottom: 8px !important; font-size: 20px !important; line-height: 30px !important;">Results</h3><div class="u-margin-s-bottom" id="sp0045" style="margin-top: 0px; margin-right: 0px; margin-left: 0px; padding: 0px; margin-bottom: 16px !important;">Extracellular cellulase production from <em style="margin: 0px; padding: 0px;">B. subtilis</em><span style="margin: 0px; padding: 0px;"><span style="margin: 0px; padding: 0px;"> strain was first confirmed on <a href="https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/carboxymethylcellulose" title="Learn more about CMC from ScienceDirect's AI-generated Topic Pages" class="topic-link" style="margin: 0px; padding: 0px; background-color: rgba(0, 0, 0, 0); word-break: break-word; text-decoration-line: underline; text-decoration-thickness: 1px; text-decoration-color: rgb(31, 31, 31); color: rgb(31, 31, 31); text-underline-offset: 1px;">CMC</a> agar and then the cellulase gene (1500 bp) was amplified from this strain and was further cloned in pET21a </span><a href="https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/expression-vector" title="Learn more about expression vector from ScienceDirect's AI-generated Topic Pages" class="topic-link" style="margin: 0px; padding: 0px; background-color: rgba(0, 0, 0, 0); word-break: break-word; text-decoration-line: underline; text-decoration-thickness: 1px; text-decoration-color: rgb(31, 31, 31); color: rgb(31, 31, 31); text-underline-offset: 1px;">expression vector</a><span style="margin: 0px; padding: 0px;">. In initial experimental studies, recombinant cellulase expression was achieved in <a href="https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/inclusion-body" title="Learn more about inclusion bodies from ScienceDirect's AI-generated Topic Pages" class="topic-link" style="margin: 0px; padding: 0px; background-color: rgba(0, 0, 0, 0); word-break: break-word; text-decoration-line: underline; text-decoration-thickness: 1px; text-decoration-color: rgb(31, 31, 31); color: rgb(31, 31, 31); text-underline-offset: 1px;">inclusion bodies</a> through shake flask level fermentation of transformed </span></span><em style="margin: 0px; padding: 0px;">E. coli</em><span style="margin: 0px; padding: 0px;"> expression host BL21DE3. Attempts were made to express this 55 KDa <a href="https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/histidine" title="Learn more about His from ScienceDirect's AI-generated Topic Pages" class="topic-link" style="margin: 0px; padding: 0px; background-color: rgba(0, 0, 0, 0); word-break: break-word; text-decoration-line: underline; text-decoration-thickness: 1px; text-decoration-color: rgb(31, 31, 31); color: rgb(31, 31, 31); text-underline-offset: 1px;">His</a><span style="margin: 0px; padding: 0px;"> tagged recombinant cellulase into soluble form by modifications in fermentation conditions. Partially purified recombinant cellulase was obtained using Ni-NTA <a href="https://www.sciencedirect.com/topics/agricultural-and-biological-sciences/affinity-chromatography" title="Learn more about affinity chromatography from ScienceDirect's AI-generated Topic Pages" class="topic-link" style="margin: 0px; padding: 0px; background-color: rgba(0, 0, 0, 0); word-break: break-word; text-decoration-line: underline; text-decoration-thickness: 1px; text-decoration-color: rgb(31, 31, 31); color: rgb(31, 31, 31); text-underline-offset: 1px;">affinity chromatography</a><span style="margin: 0px; padding: 0px;">. The activity of the purified <a href="https://www.sciencedirect.com/topics/biochemistry-genetics-and-molecular-biology/enzyme" title="Learn more about enzyme from ScienceDirect's AI-generated Topic Pages" class="topic-link" style="margin: 0px; padding: 0px; background-color: rgba(0, 0, 0, 0); word-break: break-word; text-decoration-line: underline; text-decoration-thickness: 1px; text-decoration-color: rgb(31, 31, 31); color: rgb(31, 31, 31); text-underline-offset: 1px;">enzyme</a> was confirmed by 3,5-dinitrosalicylic acid (DNS) qualitative assay.</span></span></span></div></div><div id="abs0015" style="margin: 0px; padding: 0px; color: rgb(31, 31, 31); font-family: ElsevierGulliver, Georgia, "Times New Roman", Times, STIXGeneral, "Cambria Math", "Lucida Sans Unicode", "Microsoft Sans Serif", "Segoe UI Symbol", "Arial Unicode MS", serif, sans-serif; font-size: 16px;"><h3 class="u-h4 u-margin-m-top u-margin-xs-bottom" id="sect0020" style="margin-right: 0px; margin-left: 0px; padding: 0px; margin-top: 24px !important; margin-bottom: 8px !important; font-size: 20px !important; line-height: 30px !important;">Conclusion</h3><div class="u-margin-s-bottom" id="sp0050" style="margin-top: 0px; margin-right: 0px; margin-left: 0px; padding: 0px; margin-bottom: 16px !important;">Soluble expression of active recombinant cellulase can be achieved by subtle alteration in the upstream process.</div></div>
Elsevier
December 2021
pdf
Journal of Genetic Engineering and Biotechnology
Soluble expression of recombinant active cellulase in E.coli using B.subtilis (natto strain) cellulase gene
mixed material
Elsevier