Robert P. Bywater

Current progress in Structure-Based Rational Drug Design marks a new mindset in drug discovery

The past decade has witnessed a paradigm shift in preclinical drug discovery with structure-based drug design (SBDD) making a comeback while high-throughput screening (HTS) methods have continued to generate disappointing results. There is a deficit of information between identified hits and the many criteria that must be fulfilled in parallel to convert them into preclinical candidates that have a real chance to become a drug. This gap can be bridged by investigating the interactions between the ligands and their receptors. Accurate calculations of the free energy of binding are still elusive; however progresses were made with respect to how one may deal with the versatile role of water. A corpus of knowledge combining X-ray structures, bioinformatics and molecular modeling techniques now allows drug designers to routinely produce receptor homology models of increasing quality. These models serve as a basis to establish and validate efficient rationales used to tailor and/or screen virtual libraries with enhanced chances of obtaining hits. Many case reports of successful SBDD show how synergy can be gained from the combined use of several techniques. The role of SBDD with respect to two different classes of widely investigated pharmaceutical targets: (a) protein kinases (PK) and (b) G-protein coupled receptors (GPCR) is discussed. Throughout these examples prototypical situations covering the current possibilities and limitations of SBDD are presented.

Protein folding: a problem with multiple solutions

There is continued interest in predicting the structure of proteins either at the simplest level of identifying their fold class or persevering all the way to an atomic resolution structure. Protein folding methods have become very sophisticated and many successes have been recorded with claims to have solved the native structure of the protein. But for any given protein, there may be more than one solution. Many proteins can exist in one of the other two (or more) different forms and some populate multiple metastable states. Here, the two-state case is considered and the key structural changes that take place when the protein switches from one state to the other are identified. Analysis of these results show that hydrogen bonding patterns and hydrophobic contacts vary considerably between different conformers. Contrary to what has often been assumed previously, these two types of interaction operate essentially independently of one another. Core packing is critical for proper protein structure and function and it is shown that there are considerable changes in internal cavity volumes in many cases. The way in which these switches are made is fold dependent. Considerations such as these need to be taken into account in protein structure prediction.

Drug design for ever, from hype to hope.

In its first 25xa0years JCAMD has been disseminating a large number of techniques aimed at finding better medicines faster. These include genetic algorithms, COMFA, QSAR, structure based techniques, homology modelling, high throughput screening, combichem, and dozens more that were a hype in their time and that now are just a useful addition to the drug-designers toolbox. Despite massive efforts throughout academic and industrial drug design research departments, the number of FDA-approved new molecular entities per year stagnates, and the pharmaceutical industry is reorganising accordingly. The recent spate of industrial consolidations and the concomitant move towards outsourcing of research activities requires better integration of all activities along the chain from bench to bedside. The next 25xa0years will undoubtedly show a series of translational science activities that are aimed at a better communication between all parties involved, from quantum chemistry to bedside and from academia to industry. This will above all include understanding the underlying biological problem and optimal use of all available data.

Membrane-spanning peptides and the origin of life.

An explanation is given as to why membrane-spanning peptides must have been the first information-rich molecules in the development of life. These peptides are stabilised in a lipid bilayer membrane environment and they are preferentially made from the simplest, and likewise oldest, of the amino acids that survive today. Transmembrane peptides can exercise functions that are essential for biological systems such as signal transduction and material transport across membranes. More complex peptides possessing catalytic properties could later develop on either side of the membrane as independently folding functional units formed by extension of the protruding ends of the transmembrane peptides within an aqueous environment and thereby give rise to more of the functions that are necessary for life. But the membrane was the cradle for the development of the first information-rich biomolecules.

The preferred conformation of dipeptides in the context of biosynthesis

Globular proteins are folded polypeptide structures comprising stretches of secondary structures (helical (α- or 310 helix type), polyproline helix or β-strands) interspersed by regions of less well-ordered structure (“random coil”). Protein fold prediction is a very active field impacting inte alia on protein engineering and misfolding studies. Apart from the many studies of protein refolding from the denatured state, there has been considerable interest in studying the initial formation of peptides during biosynthesis, when there are at the outset only a few residues in the emerging polypeptide. Although there have been many studies employing quantum chemical methods of the conformation of dipeptides, these have mostly been carried out in the gas phase or simulated water. None of these conditions really apply in the interior confines of the ribosome. In the present work, we are concerned with the conformation of dipeptides in this low dielectric environment. Furthermore, only the residue types glycine and alanine have been studied by previous authors, but we extend this repertoire to include leucine and isoleucine, position isomers which have very different structural propensities.

The dipeptide conformations of all twenty amino acid types in the context of biosynthesis

There have been many studies of dipeptide structure at a high level of accuracy using quantum chemical methods. Such calculations are resource-consuming (in terms of memory, CPU and other computational imperatives) which is the reason why most previous studies were restricted to the two simplest amino-acid residue types, glycine and alanine. We improve on this by extending the scope of residue types to include all 20 naturally occurring residue types. Our results reveal differences in secondary structure preferences for the all residue types. There are in most cases very deep energy troughs corresponding either to the polyproline II (collagen) helix and the α-helix or both. The β-strand was not strongly favoured energetically although the extent of this depression in the energy surface is, while not “deeper” (energetically), has a wider extent than the other two types of secondary structure. There is currently great interest in the question of cotranslational folding, the extent to which the nascent polypeptide begins to fold prior to emerging from the ribosome exit tunnel. Accordingly, while most previous quantum studies of dipeptides were carried out in the (simulated) gas or aqueous phase, we wished to consider the first step in polypeptide biosynthesis on the ribosome where neither gas nor aqueous conditions apply. We used a dielectric constant that would be compatible with the water-poor macromolecular (ribosome) environment.

Prediction of protein structural features from sequence data based on Shannon entropy and Kolmogorov complexity.

While the genome for a given organism stores the information necessary for the organism to function and flourish it is the proteins that are encoded by the genome that perhaps more than anything else characterize the phenotype for that organism. It is therefore not surprising that one of the many approaches to understanding and predicting protein folding and properties has come from genomics and more specifically from multiple sequence alignments. In this work I explore ways in which data derived from sequence alignment data can be used to investigate in a predictive way three different aspects of protein structure: secondary structures, inter-residue contacts and the dynamics of switching between different states of the protein. In particular the use of Kolmogorov complexity has identified a novel pathway towards achieving these goals.

On dating stages in prebiotic chemical evolution

The notion that RNA must have had a unique and decisive role in the development of life needs hardly be questioned. However, the chemical complexity and other properties of RNA, such as high solubility in water and vulnerability to degradation, make it improbable that RNA could have had an early presence in the development of life on Earth or on any comparable telluric planet. Rather, the task of origin of life research must surely be to identify those chemical processes which could have taken place on Earth that could accumulate the complexity and rich molecular information content needed to sustain primitive life, and ultimately give rise to RNA. A collection of likely chemical precursors to modern biomolecules is listed here together with calculations of their molecular complexity. These complexity scores are then used to propose an ordering, on a timescale, of when they might have appeared on Earth. These pre-RNA living systems would have flourished during the first ~0.3 Gyrs after the start of the Archaean era (~4.2 Gyr ago). If there ever was an “RNA-world” it could have started after that initial period (~3.9 Gyrs ago), later to be complemented with the appearance of duplex DNA at about ~3.6 Gyrs ago, some time before the earliest known stromatolites (~3.4 Gyr).

Comments on the paper "Levinthal's question revisited, and answered" by A. Ben-Naim.

The first of these two items has been dealt with at length by this author, both in terms of what is meant by a global minimum in Gibbs energy (Bywater, 2012) and in terms of where one arrives at when the Anfinsen experiment is actually carried out (Seddon & Bywater, 2012). Both of these papers, in very different ways, support the contention #1 above, and this issue will therefore not be considered further here. Disputes between competing theories about protein stability (or anything else, come to that) cannot be settled by rhetoric or invective. At some point, a set of facts has to be agreed on, and, if possible some numbers have to be produced in order to be able to compare things properly. As Lord Kelvin remarked “If you cant measure something in numbers, your knowledge of it is not really scientific”. (Well, he made some mistakes too, with his embarrassing statement about “heavier than air flying machines”, but nobody is perfect ...). At the same time, theoreticians can take some comfort from the maxim of Francis Crick to the effect that “any theory which fits all the facts must be wrong, as some of the facts themselves will be in error”. With this as a backdrop to my entry to this discussion, I wish to propose the following general guideline as to how to proceed from discord to harmony in any debate of this sort. Since the problems we are dealing with are so complex – there is no convenient “E = mc type” of solution – what we need is a set of different theoretical approaches, all of them based on sound principles of thermodynamics, mechanics, evolutionary data, etc. but which, when taken together, converge to a single (one might then say, inescapable) conclusion. In my view, we have just such a scenario here in both of the points raised in the “Levinthal revisited” paper. Having dealt with #1 (at least to my satisfaction), I proceed to comment on #2: As pointed out by Ben-Naim (2011), we have been beguiled by the very appealing arguments put forward 50 + years ago by Kauzmann (1959). At the time, it seemed convincing for several reasons.

Why twenty amino acid residue types suffice(d) to support all living systems

It is well known that proteins are built up from an alphabet of 20 different amino acid types. These suffice to enable the protein to fold into its operative form relevant to its required functional roles. For carrying out these allotted functions, there may in some cases be a need for post-translational modifications and it has been established that an additional three types of amino acid have at some point been recruited into this process. But it still remains the case that the 20 residue types referred to are the major building blocks in all terrestrial proteins, and probably universally. Given this fact, it is surprising that no satisfactory answer has been given to the two questions: why 20? and why just these 20?. Furthermore, a suggestion is made as to how these 20 map to the codon repertoire which in principle has the capacity to cater for 64 different residue types. Attempts are made in this paper to answer these questions by employing a combination of quantum chemical and chemoinformatic tools which are applied to the standard 20 amino acid types as well as 3 “non-standard” types found in nature, a set of fictitious but feasible analog structures designed to test the need for greater coverage of function space and the collection of candidate alternative structures found either on meteorites or in experiments designed to reconstruct pre-life scenarios.