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Review
. 2012 Oct 8;13(10):12857-79.
doi: 10.3390/ijms131012857.

Fragment-based screening by protein crystallography: successes and pitfalls

Zorik Chilingaryan  1 , Zhou Yin, Aaron J Oakley
Affiliations
Review

Fragment-based screening by protein crystallography: successes and pitfalls

Zorik Chilingaryan et al. Int J Mol Sci. .

Abstract

Fragment-based drug discovery (FBDD) concerns the screening of low-molecular weight compounds against macromolecular targets of clinical relevance. These compounds act as starting points for the development of drugs. FBDD has evolved and grown in popularity over the past 15 years. In this paper, the rationale and technology behind the use of X-ray crystallography in fragment based screening (FBS) will be described, including fragment library design and use of synchrotron radiation and robotics for high-throughput X-ray data collection. Some recent uses of crystallography in FBS will be described in detail, including interrogation of the drug targets β-secretase, phenylethanolamine N-methyltransferase, phosphodiesterase 4A and Hsp90. These examples provide illustrations of projects where crystallography is straightforward or difficult, and where other screening methods can help overcome the limitations of crystallography necessitated by diffraction quality.

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Figures

Figure 1
Figure 1
Structure Beta secretase-1 (BACE-1). (a) Overall fold showing location of active site; (b) Active site residues; (c) Hydroxyethylamine-based peptidomimetic inhibitor; (d) Same compound shown in BACE-1. Sub-sites are labeled according to the amino-acids either side of the cleavage site (S2, S1, S1′, S2′, etc.).
Figure 2
Figure 2
Crystal structures of fragment complexes with BACE-1. The chemical structures of the protonated forms of the compounds are shown, and, where determined, IC50 values are shown. (a) 1-aminoisoquinoline; (b) 2-aminoquinoline; (c) 2-amino-3- (benzylamino)pyridine; (d) 4-(4-fluorobenzyl)piperidine.
Figure 3
Figure 3
Chemical structures of BACE-1 inhibitors developed from fragment hits. The structural formulae of the protonated forms of the compounds bound are shown, and, where determined, IC50 values and crystal structures of complexes with BACE-1 are shown. (a) 2-amino-6-phenethylpyridine; (b) 2-(2-(1H-indol-6-yl)ethyl)-6-aminopyridine; (c) 2-amino-6-(2-(3′-methoxy-[1,1′-biphenyl]-3-yl)ethyl)pyridine; (d) 2-amino-3-((3- (pyridin-3-yl)benzyl)amino)pyridine; (e) 2-amino-3-((3-(5-methoxypyridin-3-yl)benzyl) amino)pyridine; (f) 2-amino-3-((3-(5-propyloxypyridin-3-yl)benzyl)amino)pyridine.
Figure 4
Figure 4
Chemical structures of high-affinity BACE-1 inhibitors. The structural formulae of the protonated forms of the compounds bound are shown, and, where determined, IC50 values and crystal structures of complexes with BACE-1 are shown. (a) 3-((3-(1H-indol-6- yl)benzyl)amino)-2-aminopyridine; (b) 3-((5-(1H-indol-6-yl)-2-(pyridin-2-ylmethoxy) benzyl)amino)-2-aminopyridine; (c) 2-amino-3-((2-(benzyloxy)-5-(1H-indol-6-yl)benzyl) amino)pyridine.
Figure 5
Figure 5
(a) Structure of human PDE4A in cartoon form; (b) Active-site of PDE4A in complex with inhibitor pentoxifylline; (c) Structure of pentoxifylline.
Figure 6
Figure 6
(a) Adenine component of cAMP; (b) Purine-like hits; (c) 4-[3-(methoyxphenyl)amino]-6-(methylsulfonyl)quinoline-3-carboxamide; (d) quinoline-like hits. Analogous nitrogen atoms are highlighted in blue.
Figure 7
Figure 7
(a) Resorcinol; (b) Imidazole; (c) 6-Chlorooxindole; (d) (S)-2-amino-3-(1Hinden- 3-yl)propanoic acid.
Figure 8
Figure 8
(a) PNMT with 6-chlorooxindole (6CO) and S-adenosyl-homocysteine (SAH) modelled in the active site (based on PDB entry 3KPY); (b) Same structure after reassignment of density to imidazole (IMI) and resorcinol (RCO) (based on PDB entry 4DM3). Protein backbone is shown in ribbon form, with residues shown in stick form. Ligands are drawn in stick form with carbon atoms colored orange.
Figure 9
Figure 9
(a) Hsp90 with pyrimidine fragment bound, and (b) with 4-chloro-6-(2,4- dichloro-5-(2-morpholinoethoxy)phenyl)pyrimidine-2-amine bound.
Figure 10
Figure 10
(a) Hsp90 with phenol-based fragment bound, and (b) with (2,4-dihydroxy-5- isopropylphenyl)(isoindolin-2-yl)methanone bound.
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