High resolution X-ray analyses of renin inhibitor-aspartic proteinase complexes

Foundling, Stephen I.; Cooper, J.B.; Watson, F.E.; Cleasby, Anne; Pearl, Laurence H.; Sibanda, B. L.; Hemmings, Andrew M.; Wood, Stephen P.; Blundell, T.L.; Valler, Martin J.; Norey, Christopher G.; Kay, John; Boger, Joshua; Dunn, B. M.; Leckie, Brenda J.; Jone, D. M.; Atrash, Butrus; Hallett, Allan; Szelke, M.

doi:10.1038/327349a0

Cited by 125 publications

(85 citation statements)

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“…Inhibitors bind in an extended conformation making P-sheet hydrogen bonds with the enzyme (James et al, 1982;Foundling et al, 1987;Suguna et al, 1992). This class of enzymes includes not only the pepsin-like enzymes with approxi-…”

Section: Introductionmentioning

confidence: 99%

A structural comparison of 21 inhibitor complexes of the aspartic proteinase from Endothia parasitica

Bailey

Cooper

1994

Protein Science

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The aspartic proteinases are an important family of enzymes associated with several pathological conditions such as hypertension (renin), gastric ulcers (pepsin), neoplastic disease (cathepsins D and E), and AIDS (HIV proteinase). Studies of inhibitor binding are therefore of great importance for design of novel inhibitors for potential therapeutic applications. Numerous X-ray analyses have shown that transition-state isostere inhibitors of aspartic proteinases bind in similar extended conformations in the active-site cleft of the target enzyme. Upon comparison of 21 endothiapepsin inhibitor complexes, the hydrogen bond lengths were found to be shortest where the isostere (Pl-P;) interacts with the enzyme's catalytic aspartate pair. Hydrogen bonds with good geometry also occur at P;, and more so at P,, where a conserved water molecule is involved in the interactions. Weaker interactions also occur at P2, where the side-chain conformations of the inhibitors appear to be more variable than at the more tightly held positions. At Pz and, to a lesser extent, P,, the side-chain conformations depend intriguingly on interactions with spatially adjacent inhibitor side chains, namely P; and P, , respectively. The tight binding at PI-P;, P,, and Pi is also reflected in the larger number of van der Waals contacts and the large decreases in solvent-accessible area at these positions, as well as their low temperature factors. Our analysis substantiates earlier proposals for the locations of protons in the transition-state complex. Aspartate 32 is probably ionized in the complexes, its charge being stabilized by 1, or sometimes 2, hydrogen bonds from the transition-state analogues at P I . The detailed comparison also indicates that the P, and P2 residues of substrate in the ES complex may be strained by the extensive binding interactions at P3, Pi, and Pi in a manner that would facilitate hydrolysis of the scissile peptide bond.Keywords: aspartic proteinase; inhibitor complexes; transition-state analogues Aspartic proteinases provide excellent subjects for studying molecular recognition because they bind a range of peptide substrates and inhibitors with varying degrees of specificity. Catalysis stems from 2 invariant aspartate residues, at positions 32 and 215 in porcine pepsin, which are thought to activate a bound water molecule to hydrolyze the scissile bond of the substrate. X-ray analyses have shown that these enzymes have deep and extended active-site clefts in which there are binding subsites[S&] for 9 amino acid residues of the ligand (see Davies, 1990, for review). Inhibitors bind in an extended conformation making P-sheet hydrogen bonds with the enzyme (James et al., 1982;Foundling et al., 1987;Suguna et al., 1992). This class of enzymes includes not only the pepsin-like enzymes with approxi-

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Section: Introductionmentioning

confidence: 99%

A structural comparison of 21 inhibitor complexes of the aspartic proteinase from Endothia parasitica

Bailey

Cooper

1994

Protein Science

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“…Figure 6 shows the conformations of three inhibitors of en do thia pepsin in two superpositions (26). The reduced inhibitors H-142 and H-256 superpose very well, while H-142 and L-363,564 superpose surprisingly well, especially on the C terminal side of the scissile bond region, considering that L-363,564 con tains a statine residue.…”

Section: Genj'lral Mode Of Bindingmentioning

confidence: 99%

“…Table 3 shows those inhibitors complexed with one of the fungal enzymes that have been examined by high-resolution X-ray diffraction. Some bind (54), and the L-363,564 inhibitor has a Kj to endothiapepsin of 16 nM (26,27).…”

Section: Ketone Hydratementioning

confidence: 99%

Structure and Function of the Aspartic Proteinases

Dunn¹

1991

Advances in Experimental Medicine and Biology

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“…During the past 15 years, aspartic proteinase±inhibitor complexes have been extensively studied in order to determine the catalytic mechanism of this group of enzymes (James & Sielecki, 1985;Suguna et al, 1987;Foundling, Cooper, Watson, Cleasby et al, 1987;Cooper et al, 1989;Abad-Zapatero et al, 1991;Hoover et al, 1991;Pitts et al, 1995). These results show that the inhibitors ®t into the substrate-binding cleft in extended conformations.…”

Section: Introductionmentioning

confidence: 99%

Structure of theRhizomucor mieheiaspartic proteinase complexed with the inhibitor pepstatin A at 2.7 Å resolution

Yang

Quail

1999

Acta Crystallogr D Biol Cryst

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Crystals of Rhizomucor miehei aspartic proteinase (RMP) complexed with pepstatin A grew in the orthorhombic space group P2 1 2 1 2 1 and were isomorphous to native RMP crystals. The unit-cell dimensions are a = 41.52, b = 50.82, c = 172.71 A Ê . There is one RMP±pepstatin A complex per asymmetric unit. The structure of the RMP±pepstatin A complex has been re®ned to a crystallographic R value of 19.3% and an R free value of 28.0% at 2.7 A Ê resolution. A pepstatin A molecule ®ts into the large substrate-binding cleft between the two domains of RMP in an extended conformation up to the alanine residue at the P2 H position. The dipeptide analogue statine residue at the P3 H ±P4 H position forms an inverse -turn (P3 H ± P1 H ) with the statine residue at the P1±P1 H position and its leucyl side chain binds back into the S1 H subsite. The inhibitor interacts with the residues of the substrate-binding pocket by both hydrogen bonds and hydrophobic interactions. The hydroxyl group of the statine residue at the P1±P1 H position forms hydrogen bonds with both catalytic aspartate residues (Asp38 and Asp237). This conformation mimics the expected transition state of the enzyme±substrate interaction. The binding of the inhibitor to the enzyme does not produce large distortions of the active site. No domain movement was observed compared with the native enzyme structure. However, the surface-¯ap region (residues 82±88) undergoes a conformational change, moving toward the inhibitor and becoming rigid owing to the formation of hydrogen bonds with the inhibitor. B-factor calculations of the two domains suggest that the C-terminal domain becomes more rigid in the complex than in the native structure.

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High resolution X-ray analyses of renin inhibitor-aspartic proteinase complexes

Cited by 125 publications

References 0 publications

A structural comparison of 21 inhibitor complexes of the aspartic proteinase from Endothia parasitica

A structural comparison of 21 inhibitor complexes of the aspartic proteinase from Endothia parasitica

Structure and Function of the Aspartic Proteinases

Structure of theRhizomucor mieheiaspartic proteinase complexed with the inhibitor pepstatin A at 2.7 Å resolution

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