X-ray analyses of aspartic proteinases IV

Newman, Matthew; Safro, Mark; Frazão, Carlos; Khan, Gulraiz; Zdanov, Alexander; Tickle, Ian J.; Blundell, Tom L.; Andreeva, N. S.

doi:10.1016/0022-2836(91)90934-x

Cited by 33 publications

(18 citation statements)

References 48 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The Gly residues at these positions have been suggested to eliminate steric interference with the catalytic apparatus by side chains [29]. Since rabbit cathepsin E exhibited high proteolytic activity compared to porcine pepsin A (Fig.…”

Section: Structural Features Of Procathepsin Ementioning

confidence: 99%

“…CTG GAC A I C CAG CCG CCA GCT GGG CCC CTC TGG ATC CTG GGG GAT GTC TTC ATC CGG CAG L e u AFP I I e G I n P r o Pro A l a G l y P r o l e u T r p I l e l e u G l y A s p V a l Phe I l e A r g G i n 3 5 0 3 6 0 TTC TAC TCG GTC T T T GAC CGT GGG AGC AAC CGC GTG GGG CTG GCC CCT GCA GTC CCC TAG Phe T y r S e r V a l P h e A s p A r g G l y S e r A s n A r g V a l G l y l e u A l a P r o A l a V a l P r o [29][30][31][32][33][34][35][36] and cathepsin E. Glnl was deduced to be the N-terminal residue of procathepsin E from the similarity in sequence between rabbit, guinea-pig, and human procathepsins E, since this residue was unidentified by Edman degradation, presumably because of the formation of a pyroglutamyl residue. The residue in italics, namely, Asn73, is a potential site of N-glycosylation.…”

mentioning

confidence: 99%

See 1 more Smart Citation

Rabbit procathepsin E and cathepsin E

Kageyama

1993

European Journal of Biochemistry

View full text Add to dashboard Cite

The structure of rabbit procathepsin E was determined by molecular cloning of its cDNA. The proenzyme consisted of 379 amino acids and had structural features common to human and guineapig procathepsin E species. The highly conserved tripeptide sequence at the active site of aspartic proteinases, Asp-Thr(Ser)-Gly, is, however, replaced by Asp-Thr-Val in rabbit procathepsin E. To our knowledge, this is the first case of such a variation in aspartic proteinases. The processed form, cathepsin E, hydrolyzed various biologically active peptides maximally at around pH 5. Tachykinins, such as substance P and neurokinin A, were hydrolyzed most rapidly, with specific cleavage of sequences essential for their activity. The rates of hydrolysis were several hundred-fold higher than those of cathepsin D. Furthermore, cathepsin E was able to inactivate a functional-domain peptide of fibroblast growth factor, the sequence of which resembles those of tachykinins, and it was active in the generation of functional peptides, such as endothelin and angiotensin I, from their respective precursors. Procathepsin E was detected at high levels in various fetal tissues, such as the liver, stomach and blood cells. At the adult stage, the proenzyme was detectable only in specific tissues, such as the urinary bladder, duodenum and colon. Northern-blot analysis showed similar stagespecific and tissue-specific expression of the mRNA for procathepsin E. Since tachykinins and other suited peptide substrates of cathepsin E have been shown to have mitogenic activity, (pro)cathepsin E may regulate the growth and differentiation of embryonic and fetal tissues by degrading or processing these peptides. The enzyme may also regulate the physiological activities of adult tissues which are mediated by substance P and related tachykinins.Procathepsin E is known as the precursor of cathepsin E, which is a non-secretory intracellular proteinase [I] Note. The nucleotide sequence reported in this paper has been submitted to the GenBankTMEMBL Data Bank with the accession number L08418.features are not found in other aspartic proteinases and they are correlated with the functional characteristics of procathepsin E, such as stability at alkaline pH [8] and the mechanism of activation of the proenzyme [8, 151. To confirm these unusual structural features, it seems appropriate to elucidate the structures of procathepsin E from different sources.The processed form of the enzyme, cathepsin E, is a typical endoproteinase and digests hemoglobin [5,8,9] and other protein substrates [ l , 8, 9, 161 efficiently. Cathepsin E has been suggested to play an important role in the intracellular processing or degradation of proteins, for example, in the processing of antigen in tissues associated with the immune system [17], in self-destruction of erythrocytes [lo] and the surface epithelial cells of the stomach [7]. However, it remains to be determined whether or not cathepsin E functions under physiological conditions, since the enzyme has been shown to be maximally activ...

show abstract

Section: Structural Features Of Procathepsin Ementioning

confidence: 99%

mentioning

confidence: 99%

Rabbit procathepsin E and cathepsin E

Kageyama

1993

European Journal of Biochemistry

View full text Add to dashboard Cite

show abstract

“…It was suggested also to result from the structural distortion of the flap due to crystal packing forces [4]. Actually, the conformation of the flap can depend on the crystal packing in aspartic proteinases, as it was revealed during comparison of the monoclinic and hexagonal forms of porcine pepsin crystals (Andreeva and Cooper, unpublished results).…”

Section: Introductionmentioning

confidence: 96%

“…X-ray crystallographic studies revealed that the three-dimensional structure of calf chymosin [3,4] is, in general, very close to that of porcine pepsin [5]; the outlinnings of binding pockets of the enzyme have nothing in particular to explain this difference. At the same time, an unusual feature has been observed in molecules of this enzyme: the conserved in non-viral aspartic proteinases Tyr 77 residue (Tyr 7s in porcine pepsin numbering), which usually forms the wall of the S] hydrophobic binding pocket, lies just in this S~ and partly $3 pockets (Fig.…”

Section: Introductionmentioning

confidence: 98%

Post X‐ray crystallographic studies of chymosin: the existence of two structural forms and the regulation of activity by the interaction with the histidine‐proline cluster of κ‐casein

Gustchina¹,

Румш²,

Ginodman³

et al. 1996

FEBS Letters

View full text Add to dashboard Cite

Calf chymosin molecules exist in the two alternative structural forms: the first one has S~ and $3 binding pockets occluded by its own Tyr 77 residue (the self-inhibited form); the second has these pockets free for a substrate binding (the active form). The preliminary incubation of the enzyme with a pentapeptide corresponding to the histidine-proline cluster of the specific substrate it-casein results in a 200-fold increase of the hydrolysis rate for the enzyme 'slow substrate'. The result suggests that the cluster is an allosteric effector that promotes the conversion of the enzyme into the active form. These data provide the experimental ground for the explanation of chymosin specificity towards K-casein.

show abstract

“…Mammalian and fungal enzymes have been extensively characterized, and their three-dimensional structure has been determined at high resolution (2)(3)(4)(5)(6)(7). The enzymes of this family are bilobal with a deep and extended cleft which can accommodate at least seven amino acid residues in the S 4 -S 3 Ј subsites 1 (8).…”

mentioning

confidence: 99%

Engineering of Porcine Pepsin

Shintani

Nomura

Ichishima

1997

Journal of Biological Chemistry

View full text Add to dashboard Cite

The S 1 substrate specificity of porcine pepsin has been altered to resemble that of fungal aspartic proteinase with preference for a basic amino acid residue in P 1 by site directed mutagenesis. On the basis of primary and tertiary structures of aspartic proteinases, the active site-flap mutants of porcine pepsin were constructed, which involved the replacement of Thr-77 by Asp (T77D), the insertion of Ser between Gly-78 and Ser-79 (G78(S)S79), and the double mutation (T77D/G78(S)S79). The specificities of the mutants were determined using p-nitrophenylalanine-based substrates containing a Phe or Lys residue at the P 1 position. The double mutant cleaved the Lys-Phe(4-NO 2 ) bonds, while wild-type enzyme digested other bonds. In addition, the pH dependence of hydrolysis of Lys-containing substrates by the double mutant indicates that the interactions between Asp-77 of the mutant and P 1 Lys contribute to the transition state stabilization. The double mutant was also able to activate bovine trypsinogen to trypsin by the selective cleavage of the Lys 6 -Ile 7 bond of trypsinogen. Results of this study suggest that the structure of the active site flap contributes to the S 1 substrate specificity for basic amino acid residues in aspartic proteinases.Aspartic endopeptidases (EC 3.4.23._) comprise a group of enzymes whose proteolytic activities are dependent on two aspartyl residues, Asp-32 and Asp-215, in pepsin numbering (1). Mammalian and fungal enzymes have been extensively characterized, and their three-dimensional structure has been determined at high resolution (2-7). The enzymes of this family are bilobal with a deep and extended cleft which can accommodate at least seven amino acid residues in the S 4 -S 3 Ј subsites 1 (8). The "flap," an antiparallel ␤-hairpin loop comprising residues 72 to 82 (pepsin numbering), projects across the cleft forming a channel into which a substrate binds. Although the enzymes are quite similar in their three-dimensional structures, there are drastic differences in the catalytic properties, especially in substrate specificities. There also have been many studies on the tertiary structures of aspartic proteinases with peptide-derived inhibitors (4, 5, 9 -14), and the relationship between their structures and substrate specificities has been demonstrated (4, 5, 9 -13, 15, 16). According to these reports, although the hydrogen bonding pattern between the main chain of inhibitor and enzyme is well conserved in all inhibitor-enzyme complexes, the differences in the size of subsites, of which residues make van der Waals contacts with the side chain of inhibitor, may control the substrate specificities in aspartic proteinases. Several attempts to examine the structural determinants of substrate specificities of aspartic proteinases by site-directed mutagenesis have been made recently (17-23). These provided direct evidence that the subtle differences in the structures of substrate binding sites of aspartic proteinases were sufficient to alter their substrate specificities.Aspartic pr...

show abstract

X-ray analyses of aspartic proteinases IV

Cited by 33 publications

References 48 publications

Rabbit procathepsin E and cathepsin E

Rabbit procathepsin E and cathepsin E

Post X‐ray crystallographic studies of chymosin: the existence of two structural forms and the regulation of activity by the interaction with the histidine‐proline cluster of κ‐casein

Engineering of Porcine Pepsin

Contact Info

Product

Resources

About