Electrochemical Biosensors Based on S-Layer Proteins

Abstract

Designing and development of electrochemical biosensors enable molecule sensing and quantification of biochemical compositions with multitudinous benefits such as monitoring, detection, and feedback for medical and biotechnological applications. Integrating bioinspired materials and electrochemical techniques promote specific, rapid, sensitive, and inexpensive biosensing platforms for (e.g., point-of-care testing). The selection of biomaterials to decorate a biosensor surface is a critical issue as it strongly affects selectivity and sensitivity. In this context, smart biomaterials with the intrinsic self-assemble capability like bacterial surface (S-) layer proteins are of paramount importance. Indeed, by forming a crystalline two-dimensional protein lattice on many sensors surfaces and interfaces, the S-layer lattice constitutes an immobilization matrix for small biomolecules and lipid membranes and a patterning structure with unsurpassed spatial distribution for sensing elements and bioreceptors. This review aims to highlight on exploiting S-layer proteins in biosensor technology for various applications ranging from detection of metal ions over small organic compounds to cells. Furthermore, enzymes immobilized on the S-layer proteins allow specific detection of several vital biomolecules. The special features of the S-layer protein lattice as part of the sensor architecture enhances surface functionalization and thus may feature an innovative class of electrochemical biosensors.

Keywords: S-layer proteins; biocompatible layer; bioinspired material; biosensor; self-assembly.

Figure 1

Schematic drawing (not drawn to scale) of the elements of an electrochemical biosensor. FETs: Field-Effect Transistors.

Figure 2

Schematic drawing (not drawn to scale) of an electrochemical biosensor with an S-layer lattice as intermediate layer for linking biorecognition elements to the Au (gold) electrode surfaces.

Figure 3

Transmission electron microscopy image of a freeze-etched and metal shadowed preparation of (a) an archaeal cell (from Methanocorpusuculum sinense), and (b) a bacterial cell (from Desulfotomaculum nigrificans). Bars, 200 nm. With permission from Sleytr et al. 2014 [7] (CC BY-NC-ND 3.0).

Figure 4

Cyclic voltammogram of the bare and the S-layer protein-coated gold electrode in 10 mM [Fe(CN)₆]^3−/4− containing 100 mM KCl at a scan rate of 50 mV/s.

Figure 5

The two types of electrochemical biosensors with three electrodes: reference (RE), working (WE), and counter (CE) connected to a potentiostat.

Figure 6

Direct and indirect transduction (not drawn to scale). In direct approach, the electron transfers are close to the surface, whereas in the indirect one, electron shuttles between the reaction site and the sensor surface. In the proposed model, the S-layer protein lattice constitutes an intermediate matrix. In the lipid-based biosensor (left), electrons transfer from the outer membrane to the inner membrane and vice versa via a channel protein. In the detection biosensor (middle and right), electrons transfer between the enzyme–substrate complex and cell/antibody and electrode surface, respectively. The S-layer lattice provides an immobilization matrix and ion reservoir. The pores of the S-layer lattice ensure no impact on the electron transfer. Fc: fragment crystallizable; rSbpA/ZZ: recombinant S-layer protein from Lysinibacillus sphaericus CCM 2177 with fused Fc-binding Z-domain (synthetic analog of immunoglobulin G (IgG-binding B—domain) of protein A of Staphylococcus aureus).

Figure 7

Generalized scheme of the construction of an S-layer ultrafiltration membrane (SUM)-based biosensor (not drawn to scale). S-layer carrying fragments or self-assembly products are deposited on a commercially available microfilter in a pressure-dependent process. The enzyme is deposited on the S-layer surface and chemically linked to the S-layer protein. The immobilized enzyme is contacted by a thin metal layer (PLD: pulse-laser-deposition). Finally, this composite structure is deposited with the metal layer side on the working electrode (WE) and mounted in a flow cell (RE: reference electrode, CE: counter electrode). The analyte is pumped in the flow cell. After passage across the modified SUM, the analyte reacts with the enzyme, which is detected by amperometry.