Enzymic replacement of the arginyl by a lysyl residue in the reactive site of soybean trypsin inhibitor

Sealock, Robert; Laskowski, M.

doi:10.1021/bi00837a032

Cited by 119 publications

(43 citation statements)

References 25 publications

(29 reference statements)

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“…We have tried to model the structure of the complex between proteinase K and PKI3 with this bond in the active site. This is not possible without major changes in the conformations of either protease or inhibitor, The peptide bond in SBTI which is hydrolyzed by trypsin inthe inhihition process is Arg-64-lle-65 [19]. It is structurally equivalent to Gly-66-Ala-67 in PKI3 which is localized in a loop connecting B-strands 4 and 5 at the surface of the protein and therefore easily accessible by the protease.…”

Section: Results and Discussionmentioning

confidence: 99%

The three‐dimensional structure of the bifunctional proteinase k/α‐amylase inhibitor from wheat (PKI3) at 2.5 Å resolution

et al. 1991

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Wheat germ contains an inhibitor for proicinase K, called PK13 (M 1 ∼ 19600) which simultaneously inhibits α‐amylase. PK13 was crystallized, space group P21, α = 43.02 (5) Å, n = 65.18 (7) Å, c = 32.33 (4) Å, β = 112.79 (9), X‐ray data were collected to 2.5 Å resolution, the structure solved by molecular replacement on the basis of the atomic coordinates of the homologous Erythrina caffra DE‐3 inhibitor, and refined with simulated annealing techniques with a current R‐factor of 21%. The three‐dimensional structure of PK13 is stabilized by two disulfide bridges and has a central β‐barrel with distorted β‐structure. In analogy to related inhibitors, the binding site for proteinase K is assumed to be located on the surface of the protein (amino acids residues 66–67), although the 75–76 peptide bond is cleaved upon binding.

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Section: Results and Discussionmentioning

confidence: 99%

The three‐dimensional structure of the bifunctional proteinase k/α‐amylase inhibitor from wheat (PKI3) at 2.5 Å resolution

et al. 1991

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“…If it were possible to generate des-Gly39-PCI, then incubation of this protein with CPA in the presence of various free amino acids should generate modified species of PCI, differing in residue 39. An analogous reaction has been demonstrated for enzymatic replacement of reactive site residues in soybean trypsin inhibitor (30).…”

mentioning

confidence: 94%

Structure of the potato inhibitor complex of carboxypeptidase A at 2.5-A resolution.

Rees¹,

Lipscomb²

1980

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

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The structure of the complex between the proteolytic enzyme carboxypeptidase A (peptidyl-L-amino-acid hydrolase, EC 3.4.17.1) and the 39-amino-acid carboxypeptidase A inhibitor from potatoes has been determined at 2.5-A resolution. A combination of multiple isomorphous replacement, molecular replacement, and noncrystallographic symmetry averaging techniques was used to solve the structure. The chain trace of the inhibitor and details of the binding interactions in the complex are described. A surprising aspect of the complex is that the carboxy-terminal peptide bond of the inhibitor has been hydrolyzed, and the carboxy-terminal glycine is trapped in the binding pocket of carboxypeptidase A. Consequently, the complex resembles a stage in the catalytic mechanism after hydrolysis of the peptide bond. The ring of tyrosine-248, which is known to undergo large conformational changes upon substrate binding, is in the "down" position and interacts with the inhibitor in the complex.

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“…Using such chromophoric metal atoms as active-site probes has been discussed by Vallee and Riordan (83). Replacement of amino acid residues in a polypeptide chain still is achieved most easily by isolating mutant proteins from bacteria, but Laskowski, Jr. and his colleagues at Purdue University have developed a method for enzymatically removing an essential arginine residue from the reactive site of soybean trypsin inhibitor (Kunitz) and replacing it with a lysine residue (84).…”

Section: Figure 17 the Effect Of Disulfide Bond Modification Of Turkmentioning

confidence: 99%

Chemical Modification of Proteins: An Overview

Feeney

Yamasaki

Geoghegan

1982

Advances in Chemistry

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Enzymic replacement of the arginyl by a lysyl residue in the reactive site of soybean trypsin inhibitor

Cited by 119 publications

References 25 publications

The three‐dimensional structure of the bifunctional proteinase k/α‐amylase inhibitor from wheat (PKI3) at 2.5 Å resolution

The three‐dimensional structure of the bifunctional proteinase k/α‐amylase inhibitor from wheat (PKI3) at 2.5 Å resolution

Structure of the potato inhibitor complex of carboxypeptidase A at 2.5-A resolution.

Chemical Modification of Proteins: An Overview

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