Chemical interactions enzyme reporters

Covalent attachment of enzymes to surfaces is often intuitively perceived as being more reliable than direct adsorption, but multisite physical interactions can in fact yield a comparably strong and stable union, as demonstrated by several biological examples. The biotin/streptavidin interaction requires a force of about 0.3 nN to be severed [Lee et al., 2007], and protein/protein interactions typically require 0.1 nN to break, but values over 1 nN have also been reported [Weisel et al., 2003]. These forces are comparable to those required to mpture weaker chemical bonds such as the gold-thiolate bond (1 nN for an alkanethiol, and even only 0.3 nN for a 1,3-aUcanedithiol [Langry et al., 2005]) and the poly(His)-Ni(NTA) bond (0.24 nN, [Levy and Maaloum, 2005]). 

In this chapter we have seen that enzymatic catalysis is initiated by the reversible interactions of a substrate molecule with the active site of the enzyme to form a non-covalent binary complex. The chemical transformation of the substrate to the product molecule occurs within the context of the enzyme active site subsequent to initial complex formation. We saw that the enormous rate enhancements for enzyme-catalyzed reactions are the result of specific mechanisms that enzymes use to achieve large reductions in the energy of activation associated with attainment of the reaction transition state structure. Stabilization of the reaction transition state in the context of the enzymatic reaction is the key contributor to both enzymatic rate enhancement and substrate specificity. We described several chemical strategies by which enzymes achieve this transition state stabilization. We also saw in this chapter that enzyme reactions are most commonly studied by following the kinetics of these reactions under steady state conditions. We defined three kinetic constants—kai KM, and kcJKM—that can be used to define the efficiency of enzymatic catalysis, and each reports on different portions of the enzymatic reaction pathway. Perturbations... 

The significance of monolayers to biochemistry lies in their close relation to the molecular interactions that take place at the interface between aqueous and hydrophobic systems such as cell membranes and within enzymes. The molecular alignment in monolayers is closely related to the lipid bHayer structure proposed for membranes (46). Since stereospecific chemical reactions of living organisms often take place at the cell boundaries, it is small wonder that during the past 25 years much monolayer research has been reported by biochemists, physiologists, and pharmaceutical chemists (47-51). 

Figure 1.6 shows the major types of recognition elements and transducing systems used for implementing (bio)chemical sensing. A detailed discussion is provided in other sections of this chapter and the selected examples described throughout this book. Recently, Wolfbeis reported a systematic review of recognition elements based on enzymes, ion-carriers and molecular interactions used in optical sensors [5]. 

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