Addressable self‐immobilization of lactate dehydrogenase across multiple length scales

Abstract

Successful nanobiotechnology implementation largely depends on control over the interfaces between inorganic materials and biological molecules. Controlling the orientations of biomolecules and their spatial arrangements on the surface may transform many technologies including sensors, to energy. Here, we demonstrate the self-organization of L-lactate dehydrogenase (LDH), which exhibits enhanced enzymatic activity and stability on a variety of gold surfaces ranging from nanoparticles to electrodes, by incorporating a gold-binding peptide tag (AuBP2) as the fusion partner for Bacillus stearothermophilus LDH (bsLDH). Binding kinetics and enzymatic assays verified orientation control of the enzyme on the gold surface through the genetically incorporated peptide tag. Finally, redox catalysis efficiency of the immobilized enzyme was detected using cyclic voltammetry analysis in enzyme-based biosensors for lactate detection as well as in biofuel cell energy systems as the anodic counterpart. Our results demonstrate that the LDH enzyme can be self-immobilized onto different gold substrates using the short peptide tag under a biologically friendly environment. Depending on the desired inorganic surface, the proposed peptide-mediated path could be extended to any surface to achieve single-step oriented enzyme immobilization for a wide range of applications.

Solid-binding peptides: smart tools for nanobiotechnology

“…SBPs are able to direct the immobilisation of enzymes onto nanomaterials without compromising their catalytic function [7,[19][20][21]. For example, a gold-binding peptide (GBP1) was used to direct the self-assembly of organophosphorus hydrolase (OPH) onto a gold nanoparticle-coated graphene chemosensor [20].…”

Direct bioelectrocatalysis at the interfaces by genetically engineered dehydrogenase

“…[29][30][31][32][33][34][35][36] Also, several research groups, including the authors, have demonstrated the use of a variety of solid-binding peptides as anchoring molecules onto the surfaces as well as providing functional integration between the enzymes/ proteins and specific inorganic supports. [32][33][34][35][36][37][38][39][40] The biological nature of these short peptide sequences and their vast ability to create selforganised assemblies on a surface under physiological conditions make them highly desirable when compared with their counterparts that may require higher temperatures and pH values, or other harsh reaction conditions. Among the industrially important enzymes, oxidoreductases are capable of catalysing key metabolic reactions.…”

Solid-binding peptides for immobilisation of thermostable enzymes to hydrolyse biomass polysaccharides

“…SBPs have been shown to direct the oriented immobilisation of enzymes onto solid supports with retention of native enzyme structure and catalytic activity (e.g. k cat /Km) [ 10 , 11 ]. For example, the immobilisation of alkaline phosphatase (AP) onto gold surfaces via a gold-binding peptide (GBP-1) has been reported [ 12 ].…”

“…SBPs have been used primarily as molecular tools for the functionalisation of nanomaterials and the practical application of SBP-enzyme fusions are limited currently to chemo/biosensors and bioassays that require exotic and expensive laboratory-based matrices that may not be realistic economically for large-scale biotechnological processes [ 8 , 10 , 13 ]. Silica, zeolite and other mesoporous silica-based materials are used commonly in industry as support matrices in a number of applications and the provision for bulk supply is well established.…”