Higher hit rates with heavier halogens

Halogen bonding is an esoteric type of molecular interaction. Any first-year chemistry student can tell you that halogens are electronegative. More advanced students learn that the electron density on a halogen attached to a carbon is not evenly distributed. Rather, an electron deficient region appears directly opposite the carbon bond on chlorine, bromine, and iodine atoms. This “σ-hole” can form attractive interactions with electron-rich moieties, such as backbone carbonyl atoms. These highly directional interactions can be useful alternatives to hydrogen bonds, especially since they allow a reduction in the number of hydrogen bond donors. But how to find them? This is the topic of a recent open-access paper in Frontiers in Chemistry by Frank Boeckler and collaborators at Eberhard Karls Universität Tübingen.

 

The researchers constructed a library of 191 commercially available halogen-enriched fragments (called HEFLibs), which we wrote about in 2019. Most fragments have a single halogen atom, though 15 have two of the same type (two chlorine atoms, for example). The initial publication had no screening data, but the new paper describes screening the library against four diverse proteins: the methyltransferase DOT1L, the oxygenase IDO1, and the kinases AAK1 and CAMK1G.

 

Ligand-detected STD NMR was used as the primary screen, with proteins present at 20 µM and fragments at 1 mM each in mixtures of two. Between 9 and 57 hits were found for each target, with unique hits for all the targets except DOT1L. Some fragments hit all four targets, including one similar to the "universal fragment" we highlighted here.

 

Interestingly, iodine-containing fragments gave higher hit rates than bromine-containing fragments, which in turn gave higher hit rates than chlorine-containing fragments. Specifically, 9 of 14 (64%) iodine-containing fragments hit at least one target, vs 51% and 35% for bromine- and chlorine-containing fragments.

 

To assess whether halogen bonding played a role, the researchers calculated maximum electrostatic potential (V_max) for each fragment; this is a measure of the size of the σ-hole. Fragment hits tended to have higher V_max values than non-hits.

 

One possible confounding influence is that aryl halides can react with cysteine residues in proteins, and indeed the researchers did find that some of their fragments are unstable in the presence of the cellular reducing agent glutathione.

 

To confirm the STD-NMR results with an orthogonal method, the researchers turned to isothermal titration calorimetry (ITC). Of 57 fragment-protein pairs tested, only ten gave K_D values less than 1 mM, and nine were against the kinases; there were even a couple single-digit micromolar binders for AAK1. ITC is less sensitive than NMR, so some of the other fragments may bind too weakly to fully characterize.

 

Unfortunately, crystallography has been unsuccessful so far, so it remains unclear whether any of the hits are actually making halogen bonding interactions with the proteins. Halogens are good at filling lipophilic pockets, so it is perhaps likely that less specific van der Waals interactions are the key affinity drivers. But the Boeckler group has been pursuing halogen bonding for more than a decade, so I look forward to seeing more on this topic.

 

And in the meantime, happy Pi Day!