Special Issue "Advances in the Design and Characterization of Heterogeneous Biocatalysts"

A special issue of Catalysts (ISSN 2073-4344). This special issue belongs to the section "Biocatalysis".

Deadline for manuscript submissions: 15 February 2020.

Special Issue Editors

Guest Editor
Dr. Juan M. Bolivar Website 1 Website 2 E-Mail
Department of Chemical Engineering, School of Chemical Sciences, Complutense University of Madrid, Spain
Interests: heterogeneous biocatalysts; enzyme stabilization; flow microreactor; process intensification
Guest Editor
Dr. Alexander Dennig Website 1 Website 2 E-Mail
Graz University of Technology, Institute of Biotechnology and Biochemical Engineering, Petersgasse 12/I and Austrian Centre of Industrial Biotechnology (acib), Petersgasse 14, 8010 Graz, Austria
Interests: enzymology; biocascades; reaction and protein engineering; green chemistry
Guest Editor
Dr. Javier Rocha-Martin Website 1 Website 2 Website 3 E-Mail
Institute of Catalysis and Petrochemistry, Spanish Council for Scientific Research (CSIC), Marie Curie 2, 28049 Madrid, Spain
Interests: protein immobilization; protein engineering; enzyme stabilization; multienzyme systems; cascade reactions

Special Issue Information

Dear Colleagues,

Heterogeneous biocatalysts are of great relevance in the development of modern (bio)chemical manufacturing processes. Enzyme immobilization has a long-standing tradition, with many successful examples found at both academic and industrial level. The body of knowledge is well-established and there is a broad repertoire of methodologies for immobilization, involving different chemistries and carrier materials. Fueled by the constant discovery of new enzymes/reactivities, new challenges need to be formulated for heterogeneous biocatalysts. The preparation of heterogeneous biocatalysts is not yet ready to solve emerging challenges, such as multimeric complex enzymes, the stabilization for application in nonconventional reaction media, multienzyme reaction systems, nor can it help with integrating catalysts into new reactor configurations. There is a pressing need to develop innovative concepts for the reuse of cofactors and the level of depiction for heterogeneous biocatalysts by structural features or spatiotemporal resolution of local concentration of reactants and products. To this end, a merged strategic research field can be formulated that would possibly be best described by the term “heterogeneous biocatalyst engineering”.

This Special Issue aims at hosting original contributions dealing with advances in the design, application, and characterization of immobilized biocatalysts. The main topics are:

  1. Immobilized new enzymes and biocascades of high synthetic relevance;
  2. New techniques of immobilization dealing with enzyme activity or stability;
  3. New materials for enzyme immobilization;
  4. Immobilized enzymes in new reactor concepts: e.g. flow microreactors, intensifying reactions and reactors...
  5. Advances in the characterization of immobilized enzymes.

Dr. Juan M. Bolivar
Dr. Alexander Dennig
Dr. Javier Rocha-Martin
Guest Editors

Manuscript Submission Information

Manuscripts should be submitted online at www.mdpi.com by registering and logging in to this website. Once you are registered, click here to go to the submission form. Manuscripts can be submitted until the deadline. All papers will be peer-reviewed. Accepted papers will be published continuously in the journal (as soon as accepted) and will be listed together on the special issue website. Research articles, review articles as well as short communications are invited. For planned papers, a title and short abstract (about 100 words) can be sent to the Editorial Office for announcement on this website.

Submitted manuscripts should not have been published previously, nor be under consideration for publication elsewhere (except conference proceedings papers). All manuscripts are thoroughly refereed through a single-blind peer-review process. A guide for authors and other relevant information for submission of manuscripts is available on the Instructions for Authors page. Catalysts is an international peer-reviewed open access monthly journal published by MDPI.

Please visit the Instructions for Authors page before submitting a manuscript. The Article Processing Charge (APC) for publication in this open access journal is 1600 CHF (Swiss Francs). Submitted papers should be well formatted and use good English. Authors may use MDPI's English editing service prior to publication or during author revisions.

Keywords

  • enzyme immobilization
  • enzyme stabilization
  • advanced/biohybrid materials
  • biocatalysis
  • biotechnology
  • green chemistry
  • bioreactor
  • heterogeneous catalysts

Published Papers (4 papers)

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Research

Open AccessArticle
Combi-CLEAs of Glucose Oxidase and Catalase for Conversion of Glucose to Gluconic Acid Eliminating the Hydrogen Peroxide to Maintain Enzyme Activity in a Bubble Column Reactor
Catalysts 2019, 9(8), 657; https://doi.org/10.3390/catal9080657 - 31 Jul 2019
Abstract
In this study combined cross-linked aggregates of catalase from bovine liver and glucose-oxidase from Aspergillus niger were prepared, and the effects of the precipitant and crosslinking agents, as well as the use of bovine serum albumin (BSA) as a feeder protein, on enzyme [...] Read more.
In this study combined cross-linked aggregates of catalase from bovine liver and glucose-oxidase from Aspergillus niger were prepared, and the effects of the precipitant and crosslinking agents, as well as the use of bovine serum albumin (BSA) as a feeder protein, on enzyme immobilization yield and thermal stability of both enzymes, were evaluated. Combi- crosslinking of enzyme aggregates (CLEAs) prepared using dimethoxyethane as precipitant, 25 mM glutaraldehyde and BSA/enzymes mass ratio of 5.45 (w/w), exhibited the highest enzyme activities and stabilities at 40 °C, pH 6.0, and 250 rpm for 5 h. The stability of both immobilized enzymes was fairly similar, eliminating one of the problems of enzyme coimmobilization. Combi-CLEAs were used in gluconic acid (GA) production in a bubble column reactor operated at 40 °C, pH 6.0 and 10 vvm of aeration, using 26 g L−1 glucose as the substrate. Results showed conversion of around 96% and a reaction course very similar to the same process using free enzymes. The operational half-life was 34 h, determined from kinetic profiles and the first order inactivation model. Combi-CLEAs of glucose-oxidase and catalase were shown to be a robust biocatalyst for applications in the production of gluconic acid from glucose. Full article
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Open AccessArticle
Increasing the Enzyme Loading Capacity of Porous Supports by a Layer-by-Layer Immobilization Strategy Using PEI as Glue
Catalysts 2019, 9(7), 576; https://doi.org/10.3390/catal9070576 - 29 Jun 2019
Cited by 1
Abstract
A new strategy to increase the enzyme-loading capacity of porous supports was investigated. Lipase from Pseudomonas fluorescens (PFL) was immobilized on octyl-agarose (OA) beads and treated with polyethyleneimine (PEI). Then, PFL was immobilized on the previous PFL layer. Next, the biocatalyst was coated [...] Read more.
A new strategy to increase the enzyme-loading capacity of porous supports was investigated. Lipase from Pseudomonas fluorescens (PFL) was immobilized on octyl-agarose (OA) beads and treated with polyethyleneimine (PEI). Then, PFL was immobilized on the previous PFL layer. Next, the biocatalyst was coated with PEI and a third layer of PFL was added. Sodium dodecyl sulfate polyacrylamide electrophoresis showed that the amount of PFL proportionally increased with each enzyme layer; however, the effects on biocatalyst activity were not as clear. Hydrolyzing 50 mM of triacetin at 25 °C, the activity of the three-layer biocatalyst was even lower than that of the bi-layer one; on the contrary its activity was higher when the activity was measured at 4 °C in the presence of 30% acetonitrile (that reduced the activity and thus the relevance of the substrate diffusion limitations). That is, the advantage of the multilayer formation depends on the specific activity of the enzyme and on the diffusion limitations of the substrate. When octyl agarose (OA)-PFL-PEI-PFL preparation was treated with glutaraldehyde, the activity was reduced, although the enzyme stability increased and the immobilization of the last PFL layer offered results similar to the one obtained using the three-layer preparation without glutaraldehyde modification (90%). Full article
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Open AccessArticle
Reuse of Lipase from Pseudomonas fluorescens via Its Step-by-Step Coimmobilization on Glyoxyl-Octyl Agarose Beads with Least Stable Lipases
Catalysts 2019, 9(5), 487; https://doi.org/10.3390/catal9050487 - 27 May 2019
Cited by 4
Abstract
Coimmobilization of lipases may be interesting in many uses, but this means that the stability of the least stable enzyme determines the stability of the full combilipase. Here, we propose a strategy that permits the reuse the most stable enzyme. Lecitase Ultra (LU) [...] Read more.
Coimmobilization of lipases may be interesting in many uses, but this means that the stability of the least stable enzyme determines the stability of the full combilipase. Here, we propose a strategy that permits the reuse the most stable enzyme. Lecitase Ultra (LU) (a phospholipase) and the lipases from Rhizomucor miehei (RML) and from Pseudomonas fluorescens (PFL) were immobilized on octyl agarose, and their stabilities were studied under a broad range of conditions. Immobilized PFL was found to be the most stable enzyme under all condition ranges studied. Furthermore, in many cases it maintained full activity, while the other enzymes lost more than 50% of their initial activity. To coimmobilize these enzymes without discarding fully active PFL when LU or RML had been inactivated, PFL was covalently immobilized on glyoxyl-agarose beads. After biocatalysts reduction, the other enzyme was coimmobilized just by interfacial activation. After checking that glyoxyl-octyl-PFL was stable in 4% Triton X-100, the biocatalysts of PFL coimmobilized with LU or RML were submitted to inactivation under different conditions. Then, the inactivated least stable coimmobilized enzyme was desorbed (using 4% detergent) and a new enzyme reloading (using in some instances RML and in some others employing LU) was performed. The initial activity of immobilized PFL was maintained intact for several of these cycles. This shows the great potential of this lipase coimmobilization strategy. Full article
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Open AccessArticle
Site-Specific Addressing of Particles and Coatings via Enzyme-Mediated Destabilization
Catalysts 2019, 9(4), 354; https://doi.org/10.3390/catal9040354 - 12 Apr 2019
Abstract
Enzyme mediated addressing (EMA) is a highly specific and easy-to-apply technology for direction and deposition of particles and coatings on surfaces. Key feature of this process is an enzymatic reaction in direct proximity to the surface, which induces the deposition. The technique has [...] Read more.
Enzyme mediated addressing (EMA) is a highly specific and easy-to-apply technology for direction and deposition of particles and coatings on surfaces. Key feature of this process is an enzymatic reaction in direct proximity to the surface, which induces the deposition. The technique has previously shown great success in the handling of biological particles. In this study, addressing of non-biological nanoparticles, in particular plastics and metals, is presented. The respective particles are stabilized by an amphiphilic, enzyme-degradable block copolymer, consisting of poly(ethylene glycol) and poly(caprolactone). After contact with the enzyme pseudomonas lipase, the particles are destabilized, due to the loss of the hydrophilic part of the block copolymer. The lipase is therefore immobilized on glass supports. Immobilization is performed via adsorption or covalent bonding to epoxide groups. All deposition experiments show that addressing of individual particles occurs precisely within the predefined areas of enzyme activity. Depending on the material and reaction conditions, intact nanoparticles or coatings from such can be gained. The quintessence of the study is the indifference of the EMA regarding particle materials. From this rationale, the technique offers near unlimited materials compatibility within a precise, easy-to-apply, and upscalable process. Full article
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Figure 1

Planned Papers

The below list represents only planned manuscripts. Some of these manuscripts have not been received by the Editorial Office yet. Papers submitted to MDPI journals are subject to peer-review.

1. Title: Substrate Identification Method for Protein Phosphatases Using Phosphorylation Mimic Phage Display
Authors: Takuya Yoshida, Kazuki Yamazaki, Takashi Yoneda, Atsushi Kaneko, and Yoshiro Chuman
Affiliation: Laboratory of Biological Chemistry, Department of Chemistry, Faculty of Science, Niigata University, Niigata, 950-2181, Japan
Abstract: Protein phosphorylation is the most widespread type of post-translational modification and disordered regulation of protein phosphorylation often causes serious diseases. Protein phosphatases are divided into two major groups: tyrosine (Tyr) phosphatases and serine/threonine (Ser/Thr) phosphatases. Substrate trapping mutants are frequently used to characterize Tyr phosphatases and identify their substrates; however, a rapid and simple method to identify substrates for Ser/Thr phosphatases has yet to be developed. The TFIIF-associating component of RNA polymerase II C-terminal domain (CTD) phosphatase/small CTD phosphatase (FCP/SCP) phosphatase family is one of the three Ser/Thr protein phosphatase families. Defects or control failures of these phosphatases is correlated with various diseases such as cancer and neuropathy; however, the pathogenic mechanism is not well understood. Recently we have reported that the development of a novel methodology, termed Phosphorylation Mimic Phage Display (PMPD), to identify the substrates for FCP/SCP type Ser/Thr phosphatase Scp1 using peptide phage display libraries with AlF4− [1]. Here we report that PMPD method using BeF3- instead of AlF4− to identify the novel substrate peptides against Scp1. After screening peptide phages bound to Scp1, we identified the several clones bound to Scp1 in a BeF3--dependent manner. Synthetic phospho-peptide BeM12-1 with the highest frequency showed the direct binding to the Scp1, although the binding was inhibited with the addition of BeF3- , indicating that the peptide binds to the active center of catalytic site in Scp1. Interestingly kinetics analysis revealed that BeM12-1 did not work as a substrate but a competitive inhibitor for Scp1. These data suggested that PMPD method may be applicable for the identification of both of novel substrates and inhibitors for FCP/SCP phosphatases family (Fig. 1).

text

Fig. 1. Model of Phosphorylation Mimic Phage Display (PMPD) method.

Reference

[1] K. Otsubo, T. Yoneda, A. Kaneko, S. Yagi, K. Furukawa, and Y. Chuman, Protein Pept. Lett., 2018, 25(1), 76-83.

2. Title: MODELLING OF KINETICS AND MASS TRANSFER IN LENTIL-SHAPED IMMOBILIZED BIOCATALYST PARTICLES

Authors: Milan Polakovič, Mário Miháľ, Jozef Markoš

Polyvinyl alcohol (PVA) gels belong to typical carriers for the immobilization of whole cells by entrapment. A PVA gel with a good chemical and mechanical stability, enabling high cell concentrations and good transport properties was developed by Vorlop et al. (1998a,b). A part of this invention was a manufacturing procedure of lentil-shaped gel particles named LentiKats. The same name has a Czech company who own the technology of production of these immobilized biocatalyst particles (Stloukal et al., 2007). Dozens of papers and several patents were published on different applications of LentiKats particles which typical dimensions are: the width of 3-4 mm and thickness of 0.2-0.4 mm.

None of these papers treated rigorously the effect of mass transfer resistance on the kinetics of a biocatalytic process in spite of that LentiKats have been promoted as particles diminishing the problem of mass transfer resistance. We have recently estimated the effective diffusion coefficient of sucrose from inverse size exclusion experiments when the slab geometry was intuitively considered to represent the shape of LentiKats particles (Schenkmeyerová et al., 2014)...

3. Title: Deciphering the kinetics of immobilized oxidoreductases through single-particle studies

Author: Fernando Lopez-Gallego

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