Metalloenzymes exploit catalytic function by incorporating metal ions in the active site. Metalloenzymes are ubiquitous in all enzyme families. In fact, the metal ions can be active as either redox unit or generalized Lewis acid; moreover, they can facilitate selection of stereo-products thanks to their peculiar coordination properties. Such plasticity in the reactive sites may require strict selection of the metal ion cofactors. Some enzymes are able to bind and remain active with different metal ions, while in other cases metal binding leads to attenuation or even inhibition of the catalytic function. Such phenomena are still not well understood; in order to elucidate them, it is crucial to determine the molecular details of the metal binding, as well as the electronic and reactive properties of metal-aided catalytic sites. Molecular modelling is crucial for such analyses: first of all, it can provide a highly controlled description of the metal binding sites both at the structural and electronic levels. Moreover, computational studied can screen out effects of different metal substitutions into the proteins. Such systematic analyses are difficult to perform from the experimental point of view, as loading of different metal ions during protein expression and purification may be heavily hindered by, for example, metal bioavailability or cytotoxicity, competitive binding of other ions, or metal depletion during protein purification processes.

The proposed workshop addresses current challenges in first-principles-based methods to investigate proteins that contain metal ions (metalloproteins). More specifically, we want to stimulate debate and exchange of ideas related to the functional role of metal ions in enzymes, touching problematic aspects such as electronic delocalization effects during metal recognition and binding to enzymatic pockets; structural (steric or electronic) effects regulating metalloenzyme activity and selectivity (with particular emphasis on long-range coupling to protein scaffold and solvent). The workshop will also address more technical aspects, like accuracy of electronic structure methods in describing enzyme-bound transition metal ions, and will open to perspectives on how first-principles-based methods can impact on the rational design of biomimetic metal-complexes with engineered functions, or small compounds able to block, or modulate, metalloenzymes.

This workshop is focused on the current methodological challenges that first-principles-based methods are nowadays facing when applied to metalloproteins. In particular, we will focus on the progress made toward a better description of the functional role of different metal ions in catalysing enzymatic reactions. Issues related to the description of the electronic and steric effects of metal ions such as Mg, Ca, Mn, Zn, Fe etc … – bound to enzymes – will be subject of discussions with opinion leaders in the field (see list of invited speakers). The main topics – see below – will mainly deal with the methodological advancements of computations, such as new functionals or multiscale schemes, for the treatment of metal-aided biological model systems with increased accuracy. We will also discuss how computation can impact experiments and practical applications. For example, we will examine how computational insights on metalloproteins can help the design of novel biomimetic metal-aided catalysts or potent inhibitors of pharmaceutically relevant metalloenzymes.

Scientists wishing to discuss their recent results on any of these issues, and willing to share on their novel computational QM-based methods/approaches for metal-aided chemical and biological systems, will be very welcome at this workshop.

From an organizational point of view, this workshop will be divided in five major areas related to the characterization of the structural and functional role of metal ions when bound to proteins:

1) Metal ion/protein recognition and binding
Metalloproteins contains one or more metal ions bound to the protein matrix, which incorporates the metal building ordered metal-centred structures. Each metal has one or more preferential geometry such as octahedral for Mg or tetrahedral for Zn ions. Ligands are amino acids, often Glu/Asp residues, and water molecules. These metallo-centred structures are not only necessary to maintain the correct folded structure of the protein, but are often functional, too. In fact, the presence of a metal-aided pocket is, in many cases, exploited to perform efficient catalytic reactions. Metal-bound water-protein-ligand exchange has been often experimentally reported as a key for catalysis. In this context, the discussion will be centred on the characterization of the main structural and chemical-physical properties that govern, first, the recognition of the correct metal ions by the proteins and, then, the formation of the active metal-centred pocket into the protein binding site. For example, is the conformational plasticity of the protein the key to allow the (acidic) metal ligands to coordinate the metal ion? In enzymes, is the metal recruited together with the substrate? Is the metal released together with the enzymatic products? Are computational methods able to properly describe the metal-protein interactions, discerning covalently from non-covalently bound metal ligands?

2) Electronic structure calculations for transition metals
Transition metals can be challenging for electronic structure calculations, especially when in presence of a partially filled d-shell. Most importantly, the relative energy of the d-orbitals is directly affected by the surrounding ligand field, and can dramatically change upon its geometrical distortion. Modelling transition metal complexes may require the use of electronic structure methods able to describe multiconfigurational states, reproduce the correct spin multiplicity, and properly estimate possible charge-delocalization over tightly bound ligands.[14] In particular, a wide range of approaches, including DFT and related protocols (spin projections, spin flipping) as well as post-HF methods (such as CAS-SCF) will be discussed.

3) Steric vs electronic effects of catalytic metal ions during catalysis
Quantum enzymology, which dissects energetics and structural features of enzymatic mechanisms, is of great interest to the scientific community. For example, insights from QM-based studies of an enzyme can be of help in the rational design of enzyme-mimetic systems and synthetic catalysts. Or, those insights can inspire the design of a small molecule able to block, or modulate, the enzymatic function. In this scenario, we will focus on the current challenges of first-principles based methods in the study of metalloproteins, which are estimated to cover between one third and a half of the entire proteome of living organisms. For example, there is evidence showing that electronic and steric effects can be used to explain the different catalytic efficiency of the same enzyme, in the presence o different catalytic metal ions.[4, 6-8] Is it really possible to discern steric and electronic effects of metal ions during catalysis? How to use QM-based insights to make informative choices on which metal ion, in a certain enzyme pocket, is more efficient and/or selective? Can we explain, through the use of QM-based methods, the different catalytic efficiency of “similar” metal ions such as Mg, Ca, Zn or Mn, bound to the same enzyme? Can we improve the description of charge localization effects in metal-aided catalytic sites? And what is the exact role of charge transfer effects for efficient catalysis? These, and other related issues dealing with steric and electronic effects of catalytic metal ions for catalysis, will be addressed in this section of the workshop.

4) Electron transfer in metalloproteins
A current challenge of major interest is the computational investigation of charge transport through biological systems, which is key for life-sustaining processes such as respiration and photosynthesis. Metalloproteins are key players in such processes, being able to fine-tuning their redox potential, and providing natural binding centres for the moving electrons (i.e. cytochromes, blue single copper proteins, etc). All these phenomena can be investigated by means of first-principles based methods, which can elucidate the mechanism used to efficiently catalyse such processes.[15-17] In this workshop, we will promote discussion on this topic, fostering both computational and experimental contributions that will clarify how theoretical methods can now be more efficiently integrated with experiments. We will in particular welcome discussions on computational characterization of redox states, computation of redox properties, and advanced simulations of electron transfer processes.

5) Biomimetic metallo-organic compounds
The chemical activity of natural molecules like a metalloenzyme can be of inspiration to design new synthetic compounds with tailored functions to be used for the most diverse technological applications. During such processes, it is most crucial to understand how the template natural system really works, and what are the minimal indispensable structural components that must be kept or mimicked in order to retain the function of interest. Most importantly, subtle structural variations can influence dramatically the reactivity, the selectivity, and the efficiency of designed compounds.[18] Given the intrinsic complexity in the wet synthesis of new molecules, molecular modelling can be of greater help in driving such process.19 QM-based and mixed QM/MM are proving of effective success in such task. Our workshop will therefore also discuss the state of the art in this field.