dc.contributor.author | Kulsharova, Gulsim | |
dc.contributor.author | Dimov, Nikolay | |
dc.contributor.author | Marques, Marco P.C. | |
dc.contributor.author | Szita, Nicolas | |
dc.contributor.author | Baganz, Frank | |
dc.date.accessioned | 2018-09-25T09:13:25Z | |
dc.date.available | 2018-09-25T09:13:25Z | |
dc.date.issued | 2017-12-11 | |
dc.identifier.citation | Kulsharova , G , Dimov , N , Marques , M P C , Szita , N & Baganz , F 2017 , ' Simplified immobilisation method for histidine-tagged enzymes in poly(methyl methacrylate) microfluidic devices ' , New Biotechnology . https://doi.org/10.1016/j.nbt.2017.12.004 | |
dc.identifier.issn | 1871-6784 | |
dc.identifier.other | ORCID: /0000-0002-2873-1505/work/63687438 | |
dc.identifier.uri | http://hdl.handle.net/2299/20644 | |
dc.description | Article in press. Kulsharova, G., New BIOTECHNOLOGY (2017), https://doi.org/10.1016/j.nbt.2017.12.004 | |
dc.description.abstract | Poly(methyl methacrylate) (PMMA) microfluidic devices have become promising platforms for a wide range of applications. Here we report a simple method for immobilising histidine-tagged enzymes suitable for PMMA microfluidic devices. The 1-step-immobilisation described is based on the affinity of the His-tag/Ni-NTA interaction and does not require prior amination of the PMMA surface, unlike many existing protocols. We compared it with a 3-step immobilisation protocol involving amination of PMMA and linking NTA via a glutaraldehyde cross-linker. These methods were applied to immobilise transketolase (TK) in PMMA microfluidic devices. Binding efficiency studies showed that about 15% of the supplied TK was bound using the 1-step method and about 26% of the enzyme was bound by the 3-step method. However, the TK-catalysed reaction producing l-erythrulose performed in microfluidic devices showed that specific activity of TK in the device utilising the 1-step immobilisation method was approximately 30% higher than that of its counterpart. Reusability of the microfluidic device produced via the 1-step method was tested for three cycles of enzymatic reaction and at least 85% of the initial productivity was maintained. The device could be operated for up to 40 h in a continuous flow and on average 70% of the initial productivity was maintained. The simplified immobilisation method required fewer chemicals and less time for preparation of the immobilised microfluidic device compared to the 3-step method while achieving higher specific enzyme activity. The method represents a promising approach for the development of immobilised enzymatic microfluidic devices and could potentially be applied to combine protein purification with immobilisation. | en |
dc.format.extent | 391567 | |
dc.format.extent | 774460 | |
dc.language.iso | eng | |
dc.relation.ispartof | New Biotechnology | |
dc.subject | Enzyme immobilisation | |
dc.subject | Histidine-tagged enzyme | |
dc.subject | Microfluidics | |
dc.subject | Poly(methyl methacrylate) (PMMA) | |
dc.subject | Transketolase | |
dc.subject | Biotechnology | |
dc.subject | Bioengineering | |
dc.subject | Molecular Biology | |
dc.title | Simplified immobilisation method for histidine-tagged enzymes in poly(methyl methacrylate) microfluidic devices | en |
dc.contributor.institution | School of Physics, Engineering & Computer Science | |
dc.description.status | Peer reviewed | |
dc.identifier.url | http://www.scopus.com/inward/record.url?scp=85038021922&partnerID=8YFLogxK | |
rioxxterms.versionofrecord | 10.1016/j.nbt.2017.12.004 | |
rioxxterms.type | Journal Article/Review | |
herts.preservation.rarelyaccessed | true | |