Biosensors and the search for structure–activity correlations

Most molecular biologists believe that the behaviour of biological macromolecules can be fully explained from their structure and they adhere wholeheartedly to the dictum: ‘Structure determines function’. Explanations in molecular biology tend to take the form of an apparently causal, linear pathway that goes from the nucleotide sequence to the protein sequence (step 1), then to the protein tertiary structure (step 2), to specific binding activity (step 3) and finally to biological function (step 4). Such a pathway seems to possess explanatory power partly because it is presented in a context-independent manner, i.e. as if the context and the cellular environment did not matter. For example, in step 1 of the pathway, the role played by the cellular machinery in bringing about the translation process is not considered. In the case of step 2, the implicit assumption is made that the intractable protein folding problem has been solved and that the physico-chemical environment during the folding process is irrelevant; furthermore the need for chaperones is also ignored. The fallacy of such assumptions was clearly demonstrated in the case of cyclosporin, which was found, rather unexpectedly, to have completely different conformations in different environments (Zeder-Lutz et al., 1993a; Altschuhet al., 1994). When it comes to step 3 in the explanatory pathway, the hypothesized determinism is contradicted by the multiplicity of structures that are compatible with the same activity and by the fact that a single structure can be linked to a variety of activities (Martinet al., 1998). Step 3 also does not take into account the fact that binding sites are relational entities defined by the interacting partners and not by intrinsic structural features that could be recognized independently of the interaction with a particular ligand. Clearly, the structure of one of the interacting molecules on its own cannot be used to deduce binding activity without considering a particular relation with a specific partner. Since a binding site cannot be visualized in the absence of a specific interaction, structure and activity must be viewed in an integrated manner and analyzed in a four-dimensional spatio-temporal framework (Van Regenmortel, 1996). Finally, step 4 adds very little in the way of explanation, partly because of the ambiguity of the term ‘function’ (Kitcher, 1998; Van Regenmortel, 1999). The mistaken belief that structure causes activity arises from the failure to realize that causality is a relation between successive events and not between material objects. A cause is an event that is a necessary and sufficient condition for the occurrence of a later event, called the effect. A biological event such as a binding reaction can thus not be caused by something that is not an event, i.e. the structure of a molecule. When attempts are made to analyse structure– function relationships, correlations rather than causal relations should be investigated. It is not possible to deduce what a biomolecule is able to do from what it looks like, the reason being that its activity depends on the context, such as the physico-chemical and cellular environment and the existence of a relational nexus with a particular ligand. Instead of trying to discover a single non-existing causal link between the structure and activity of a biomolecule, the aim should be to uncover correlations between the atomic structure of binding sites and biological activity and to analyse the multiplicity of factors that can influence the activity. Since the primary requirement for biological activity is binding, the advent of biosensor instruments that measure binding kinetics with considerable ease and precision has greatly simplified the search for structure–activity correlations in biomolecules (Karlsson and Roos, 1997; Malmqvist and Karlsson, 1997; Myszka, 1997; Schuck, 1997; Fivash et al., 1998). The most successful biosensor instruments are based on the optical phenomenon of surface plasmon resonance and they allow the progress of binding to be followed in real time without the need to label the reactants. Biosensors make it possible to find out which molecules interact, how many of them interact, how quickly and how strongly they interact, what the binding stoichiometry is and whether conformational changes occur upon binding. Sitedirected mutagenesis and alanine-scanning mutagenesis studies make it possible to measure the change in binding characteristics resulting from each amino acid residue substitution and to evaluate the energetic contribution of individual side-chains to the binding interaction (Malmqvist, 1996; Kelley, 1996). The high reproducibility of biosensor data allows 10–20% differences in dissociation rate constants to be detected (Rauffer-Bruye `re et al., 1997) and this provides the most detailed information available so far for correlating the atomic coordinates of a protein–protein complex with the binding energy and biological activity. During the last 10 years, theJournal of Molecular Recognitionhas published many papers describing the use of biosensor technology for analyzing biochemical interactions (Fägerstamet al., 1990; Margulieset al., 1993; ZederLutz et al., 1993b; Mortonet al., 1994; Jendeberget al., 1995; Johnsson et al., 1995; Okazaki et al., 1995; Chatellier et al., 1996; Doyleet al., 1997; Korttet al., 1997; Edwards et al., 1997; Ohlsonet al., 1997; Nilssonet al., 1997; Roos et al., 1998; Van Regenmortel et al., 1998; Nelson and Krone, 1999; Evans and MacKenzie, 1999; Huber et al., 1999). In view of this trend, it was decided that JMR will publish a yearly review summarizing major advances in the use of biosensors for analyzing molecular recognition. The first such review covering papers published in 1998 has been prepared by D.G. Myszka and will appear in a forthcoming issue. The present issue of the journal contains a number of review papers presented at a recent INSERM Workshop held in Paris, 7–8 June 1999, which discussed the use of JOURNAL OF MOLECULAR RECOGNITION J. Mol. Recognit.1999;12:277–278