Synthesis of analogs of the carboxyl protease inhibitor pepstatin. Effect of structure on inhibition of pepsin and renin

Rich, Daniel H.; Sun, Eric T.; Ulm, Edgar H.

doi:10.1021/jm00175a006

Cited by 134 publications

(45 citation statements)

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“…Unfortunately, an Ackermann-Potter plot was not per formed at that time [1], From the present work, however, it is clear that, like the situa tion with renin and cathepsin-D, pepstatin inhibition of pepsin is also of the tight-bind ing type. These results with pepsin are in agreement with other recent studies [10,11].…”

Section: Inhibition Of Pepsin By Pepstatinsupporting

confidence: 83%

Pepstatin Inhibition of Proteinase-Free Human Renin

Pourmotabbed¹,

Chou²,

Gregerman³

1982

Enzyme

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The kinetics of inhibition of proteinase-free human renin and of pepsin by the carboxyl proteinase inhibitor pepstatin have been examined. Inhibition was of the tightbinding type with both enzymes. Inhibition of crude human renin was of the classical, freely reversible type, but most of the renin-like activity of the preparation was due to contaminating proteinases which could be separated chromatographically from the renin. These results clarify previous kinetic studies of pepstatin inhibition of human renin.

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Section: Inhibition Of Pepsin By Pepstatinsupporting

confidence: 83%

Pepstatin Inhibition of Proteinase-Free Human Renin

Pourmotabbed¹,

Chou²,

Gregerman³

1982

Enzyme

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“…The stereochemical configuration of the 4-hydroxyl group in the hydroxyethylene isosteres is crucial for potent inhibition: compound 7 possessing a (4R)-hydroxyl group is 80 times less potent than its (4S)-diastereomer (compound 4). Preferential binding of the (S)-hydroxyl stereoisomer is a general phenomenon observed with both hydroxyethylene isostere and statine analogue inhibitors of other aspartic proteases as well (20,33,37).…”

Section: Resultsmentioning

confidence: 99%

“…The other inhibitors in Table 2 Table 2); Cbz is benzyloxycarbonyl. Compounds 19,27, and 30 were prepared using published procedures (20)(21)(22). The synthesis of compounds [20][21][22][23][24][25][26]28, and 29 will be described elsewhere (G.B.D., unpublished data).…”

Section: Methodsmentioning

confidence: 99%

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Inhibition of human immunodeficiency virus 1 protease in vitro: rational design of substrate analogue inhibitors.

Dreyer¹,

Metcalf²,

Tomaszek³

et al. 1989

Proc. Natl. Acad. Sci. U.S.A.

193

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Inhibitors of the protease from human immunodeficiency virus 1 (HIV-1) were designed, synthesized, and kinetically characterized. Analogues of a heptapeptide substrate of HIV-1 protease with sequence similar to the pl7-p24 cleavage site in the natural substrate, Pr55519, were synthesized in which the scissile dipeptide bond was replaced with bonds from six categories of stable mimics of an aspartic proteolysis transition state or intermediate. These mimics included an analogue of statine, hydroxyethylene isosteres, two categories of phosphinic acids, a reduced amide isostere, and an a,a-difluoroketone. The resulting peptide analogues were linear competitive inhibitors of purified recombinant HIV-1 protease with inhibition constants ranging from 18 nM to 40 ,uM depending on the type of inhibitor. A truncated inhibitor, an analogue of a hexapeptide, retained full inhibitory potency. The most potent inhibitors, containing the hydroxyethylene isostere, effectively blocked the proteolytic processing of a recombinant form of Pr55s'9 by HIV-1 protease in a cell-free assay.

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“…t-Butoxycarbonyl (Boc)-StaOEt (12) was deprotected with 4 M HCl in dioxane and sequentially coupled with Boc-L-valine, Boc-L-valine, and isovaleryl anhydride by using procedures previously described for-the synthesis of pepstatin analogues (13) (17) for two reflections. The falloff of intensity of the diffraction pattern due to radiation damage was corrected by a function of time and 0 (18).…”

Section: Methodsmentioning

confidence: 99%

Conformational flexibility in the active sites of aspartyl proteinases revealed by a pepstatin fragment binding to penicillopepsin.

James¹,

Sielecki²,

Salituro³

et al. 1982

Proc. Natl. Acad. Sci. U.S.A.

158

123

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BiochemistryConformational flexibility in the active sites of aspartyl proteinases revealed by a pepstatin fragment binding to penicillopepsin ( Communicated by Joseph S. Fruton, June 21, 1982 ABSTRACT Crystals of the molecular complex between the esterified tripeptide fragment of pepstatin and the aspartyl proteinase penicillopepsin are isomorphous with crystals of native penicillopepsin. The difference electron-density map at 1.8-A resolution, computed by using the amplitude differences and refined phases of reflections from the crystal of native penicillopepsin, unambiguously showed the binding mode of isovaleryl Pepstatin is a naturally occurring inhibitor of aspartyl proteinases (1); it has the amino acid composition isovaleryl (Iva)-ValVal-Sta-Ala-StaOH, in which Sta is the residue of statine [(4S,3S)-4-amino-3-hydroxyl-6-methylheptanoic acid] (2). The interaction ofpepstatin with aspartyl proteases is characterized by small dissociation constants [i.e., 4.6 x 10-11 M with porcine pepsin (3) and 1.5 x 10-10 M with penicillopepsin (unpublished results)]. It Structure I Penicillopepsin is an aspartyl proteinase isolated from the mold Penicillium janthinellum. Its crystal structure (6, 7) has been refined at 1.8-A resolution (8) to a conventional R-factor § of 0.136 for those reflections with I, the intensity of Bragg reflections, equal to or greater than the estimated standard deviation of the intensity measurement derived from counting statistics [I 2 o-(I)]. The aspartyl proteinases are characterized by a long binding cleft that can accommodate 7 or 8 amino acid residues of an oligopeptide substrate in an extended conformation. The two catalytically important aspartyl residues, Asp&jand Asp-213, are centrally located in this binding cleft (Asp32and Asp-215 in the porcine pepsin sequence numbering).Fluorescence measurements on the aspartyl proteinases with specific substrates have indicated that conformational mobility of groups in the active site may play an important role in the mechanism of these enzymes (9). MATERIALS AND METHODSThe enzyme, penicillopepsin, was prepared, purified, and crystallized as described (6,7,10). Crystals of the native enzyme The pepstatin derivative Iva-Val-Val-StaOEt was synthesized by the following procedure. t-Butoxycarbonyl (Boc)-StaOEt (12) was deprotected with 4 M HCl in dioxane and sequentially coupled with Boc-L-valine, Boc-L-valine, and isovaleryl anhydride by using procedures previously described for-the synthesis of pepstatin analogues (13) 6137The publication costs ofthis article were defrayed in part by page charge payment. This article must therefore be hereby marked "advertisement" in accordance with 18 U. S. C. §1734 solely to indicate this fact.

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Synthesis of analogs of the carboxyl protease inhibitor pepstatin. Effect of structure on inhibition of pepsin and renin

Cited by 134 publications

References 1 publication

Pepstatin Inhibition of Proteinase-Free Human Renin

Pepstatin Inhibition of Proteinase-Free Human Renin

Inhibition of human immunodeficiency virus 1 protease in vitro: rational design of substrate analogue inhibitors.

Conformational flexibility in the active sites of aspartyl proteinases revealed by a pepstatin fragment binding to penicillopepsin.

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