DOI: 10.1111/j.1432-1033.1973.tb02660.x. On the Mode of Activation of the Catalytically Essential Sulfhydryl Group of Papain

On the Mode of Activation of the Catalytically Essential Sulfhydryl Group of Papain

10.1111/j.1432-1033.1973.tb02660.x

Crossref journal-article

Wiley

European Journal of Biochemistry (311)

Abstract

The reaction of the thiol group of papain with chloroacetamide, iodoacetamide, d‐ and l‐2‐bromopropionamides was studied. In the acidic pH‐range the reaction rate is higher than expected for an ordinary SH‐group and shows a double‐sigmoid pH‐rate profile. In addition to the already described pKa of 8.5, we found a pKa of 4.0. This indicates that in the pH‐range where the enzyme is catalytically active the thiol group of Cys‐25 interacts with some amino acid side chain, presumably with the imidazole group of the neighboring His‐159. In the light of the X‐ray diffraction studies (Drenth et al., 1971), it is probable that the anomalous PKa of His‐159 is due to its interaction with Trp‐177.The comparison of the pH‐rate profiles of d‐ and l‐2‐bromopropionamide reactions indicates that there is no sifgnificant alteration in the geometry of the active site around pH 4, whereas at slightly alkaline pH structural changes can be observed. 2H2O has no effect on the rate constants of the alkylation reaction. This rules out the possibility of general base catalysis by the imidazole group of His‐159.The experimental data indicate that the dissociated thiol group forms an ion pair with the protonated imidazole group. On the basis of the formation of thiolate‐imidazolium ion pair, the mechanism of action of papain is reconsidered. It is concluded that the acylation and deacylation steps are not symmetrical processes, and they are one‐encounter type reactions. The interaction between His‐159 and Trp‐177 may be important in the catalysis.

Bibliography

PolgÃ¡r, L. (1973). On the Mode of Activation of the Catalytically Essential Sulfhydryl Group of Papain. European Journal of Biochemistry, 33(1), 104â109. Portico.

Authors 1

László Polgár (first)

References 24 Referenced 84

10.1016/S0021-9258(18)63630-0 / J. Biol. Chem. by Chaikin I. M. (1969)
10.1111/j.1432-1033.1968.tb19526.x
{'key': 'e_1_2_3_4_2', 'first-page': '502', 'volume-title': 'The Enzymes', 'author': 'Glazer A. N.', 'year': '1971'} / The Enzymes by Glazer A. N. (1971)
10.1016/0003-9861(59)90090-6
10.1021/j100830a521
{'key': 'e_1_2_3_7_2', 'first-page': '3', 'volume-title': 'Mechanism and Control Properties of Phosphotransferases', 'author': 'Polák L.', 'year': '1971'} / Mechanism and Control Properties of Phosphotransferases by Polák L. (1971)
10.1111/j.1432-1033.1968.tb00371.x
10.1021/bi00840a067
10.1016/S0065-3233(08)60279-X
{'key': 'e_1_2_3_11_2', 'first-page': '36', 'volume': '31', 'author': 'Lowe G.', 'year': '1970', 'journal-title': 'Biochem. Soc. Symp.'} / Biochem. Soc. Symp. by Lowe G. (1970)
{'key': 'e_1_2_3_12_2', 'first-page': '484', 'volume-title': 'The Enzymes', 'author': 'Drenth J.', 'year': '1971'} / The Enzymes by Drenth J. (1971)
10.1021/ja00976a033
10.1021/bi00866a053
10.1042/bj1080855
10.1021/bi00854a032
10.1016/0005-2795(70)90123-6
10.1016/0003-9861(66)90039-7
{'key': 'e_1_2_3_19_2', 'first-page': '29', 'volume': '7', 'author': 'Polgár L.', 'year': '1972', 'journal-title': 'Acta Biochim. Biophys. Acad. Sci. Hung.'} / Acta Biochim. Biophys. Acad. Sci. Hung. by Polgár L. (1972)
10.1098/rstb.1970.0023
10.1042/bj1240117
{'key': 'e_1_2_3_22_2', 'volume': '7', 'author': 'Polgár L.', 'year': '1972', 'journal-title': 'Acta Biochim. Biophys. Acad. Sci. Hung.'} / Acta Biochim. Biophys. Acad. Sci. Hung. by Polgár L. (1972)
10.1021/ja01090a034
10.1073/pnas.47.12.1924
10.1021/ja00872a024

Dates

Type	When
Created	20 years, 5 months ago (March 4, 2005, 10:01 a.m.)
Deposited	1 year, 9 months ago (Nov. 23, 2023, 10:06 a.m.)
Indexed	4 months, 3 weeks ago (April 2, 2025, 4:24 a.m.)
Issued	52 years, 6 months ago (Feb. 1, 1973)
Published	52 years, 6 months ago (Feb. 1, 1973)
Published Online	20 years, 5 months ago (March 4, 2005)
Published Print	52 years, 6 months ago (Feb. 1, 1973)

Funders 0

None

BibTeX

@article{Polg_r_1973, title={On the Mode of Activation of the Catalytically Essential Sulfhydryl Group of Papain}, volume={33}, ISSN={1432-1033}, url={http://dx.doi.org/10.1111/j.1432-1033.1973.tb02660.x}, DOI={10.1111/j.1432-1033.1973.tb02660.x}, number={1}, journal={European Journal of Biochemistry}, publisher={Wiley}, author={Polgár, László}, year={1973}, month=feb, pages={104–109} }

JSON

{
  "indexed": {
    "date-parts": [
      [
        2025,
        4,
        2
      ]
    ],
    "date-time": "2025-04-02T08:24:44Z",
    "timestamp": 1743582284185
  },
  "reference-count": 24,
  "publisher": "Wiley",
  "issue": "1",
  "license": [
    {
      "start": {
        "date-parts": [
          [
            2005,
            3,
            4
          ]
        ],
        "date-time": "2005-03-04T00:00:00Z",
        "timestamp": 1109894400000
      },
      "content-version": "vor",
      "delay-in-days": 11719,
      "URL": "http://onlinelibrary.wiley.com/termsAndConditions#vor"
    }
  ],
  "content-domain": {
    "domain": [],
    "crossmark-restriction": false
  },
  "published-print": {
    "date-parts": [
      [
        1973,
        2
      ]
    ]
  },
  "abstract": "<jats:p>The reaction of the thiol group of papain with chloroacetamide, iodoacetamide, <jats:sc>d</jats:sc>\u2010 and <jats:sc>l</jats:sc>\u20102\u2010bromopropionamides was studied. In the acidic pH\u2010range the reaction rate is higher than expected for an ordinary SH\u2010group and shows a double\u2010sigmoid pH\u2010rate profile. In addition to the already described p<jats:italic>K</jats:italic><jats:sub>a</jats:sub> of 8.5, we found a p<jats:italic>K</jats:italic><jats:sub>a</jats:sub> of 4.0. This indicates that in the pH\u2010range where the enzyme is catalytically active the thiol group of Cys\u201025 interacts with some amino acid side chain, presumably with the imidazole group of the neighboring His\u2010159. In the light of the X\u2010ray diffraction studies (Drenth <jats:italic>et al.</jats:italic>, 1971), it is probable that the anomalous P<jats:italic>K</jats:italic><jats:sub>a</jats:sub> of His\u2010159 is due to its interaction with Trp\u2010177.</jats:p><jats:p>The comparison of the pH\u2010rate profiles of <jats:sc>d</jats:sc>\u2010 and <jats:sc>l</jats:sc>\u20102\u2010bromopropionamide reactions indicates that there is no sifgnificant alteration in the geometry of the active site around pH 4, whereas at slightly alkaline pH structural changes can be observed.</jats:p><jats:p> <jats:sup>2</jats:sup>H<jats:sub>2</jats:sub>O has no effect on the rate constants of the alkylation reaction. This rules out the possibility of general base catalysis by the imidazole group of His\u2010159.</jats:p><jats:p>The experimental data indicate that the dissociated thiol group forms an ion pair with the protonated imidazole group. On the basis of the formation of thiolate\u2010imidazolium ion pair, the mechanism of action of papain is reconsidered. It is concluded that the acylation and deacylation steps are not symmetrical processes, and they are one\u2010encounter type reactions. The interaction between His\u2010159 and Trp\u2010177 may be important in the catalysis.</jats:p>",
  "DOI": "10.1111/j.1432-1033.1973.tb02660.x",
  "type": "journal-article",
  "created": {
    "date-parts": [
      [
        2005,
        3,
        4
      ]
    ],
    "date-time": "2005-03-04T15:01:38Z",
    "timestamp": 1109948498000
  },
  "page": "104-109",
  "source": "Crossref",
  "is-referenced-by-count": 84,
  "title": "On the Mode of Activation of the Catalytically Essential Sulfhydryl Group of Papain",
  "prefix": "10.1111",
  "volume": "33",
  "author": [
    {
      "given": "L\u00e1szl\u00f3",
      "family": "Polg\u00e1r",
      "sequence": "first",
      "affiliation": []
    }
  ],
  "member": "311",
  "published-online": {
    "date-parts": [
      [
        2005,
        3,
        4
      ]
    ]
  },
  "reference": [
    {
      "key": "e_1_2_3_2_2",
      "doi-asserted-by": "crossref",
      "first-page": "5087",
      "DOI": "10.1016/S0021-9258(18)63630-0",
      "volume": "244",
      "author": "Chaikin I. M.",
      "year": "1969",
      "journal-title": "J. Biol. Chem."
    },
    {
      "key": "e_1_2_3_3_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1111/j.1432-1033.1968.tb19526.x"
    },
    {
      "key": "e_1_2_3_4_2",
      "first-page": "502",
      "volume-title": "The Enzymes",
      "author": "Glazer A. N.",
      "year": "1971"
    },
    {
      "key": "e_1_2_3_5_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1016/0003-9861(59)90090-6"
    },
    {
      "key": "e_1_2_3_6_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1021/j100830a521"
    },
    {
      "key": "e_1_2_3_7_2",
      "first-page": "3",
      "volume-title": "Mechanism and Control Properties of Phosphotransferases",
      "author": "Pol\u00e1k L.",
      "year": "1971"
    },
    {
      "key": "e_1_2_3_8_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1111/j.1432-1033.1968.tb00371.x"
    },
    {
      "key": "e_1_2_3_9_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1021/bi00840a067"
    },
    {
      "key": "e_1_2_3_10_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1016/S0065-3233(08)60279-X"
    },
    {
      "key": "e_1_2_3_11_2",
      "first-page": "36",
      "volume": "31",
      "author": "Lowe G.",
      "year": "1970",
      "journal-title": "Biochem. Soc. Symp."
    },
    {
      "key": "e_1_2_3_12_2",
      "first-page": "484",
      "volume-title": "The Enzymes",
      "author": "Drenth J.",
      "year": "1971"
    },
    {
      "key": "e_1_2_3_13_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1021/ja00976a033"
    },
    {
      "key": "e_1_2_3_14_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1021/bi00866a053"
    },
    {
      "key": "e_1_2_3_15_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1042/bj1080855"
    },
    {
      "key": "e_1_2_3_16_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1021/bi00854a032"
    },
    {
      "key": "e_1_2_3_17_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1016/0005-2795(70)90123-6"
    },
    {
      "key": "e_1_2_3_18_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1016/0003-9861(66)90039-7"
    },
    {
      "key": "e_1_2_3_19_2",
      "first-page": "29",
      "volume": "7",
      "author": "Polg\u00e1r L.",
      "year": "1972",
      "journal-title": "Acta Biochim. Biophys. Acad. Sci. Hung."
    },
    {
      "key": "e_1_2_3_20_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1098/rstb.1970.0023"
    },
    {
      "key": "e_1_2_3_21_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1042/bj1240117"
    },
    {
      "key": "e_1_2_3_22_2",
      "volume": "7",
      "author": "Polg\u00e1r L.",
      "year": "1972",
      "journal-title": "Acta Biochim. Biophys. Acad. Sci. Hung."
    },
    {
      "key": "e_1_2_3_23_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1021/ja01090a034"
    },
    {
      "key": "e_1_2_3_24_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1073/pnas.47.12.1924"
    },
    {
      "key": "e_1_2_3_25_2",
      "doi-asserted-by": "publisher",
      "DOI": "10.1021/ja00872a024"
    }
  ],
  "container-title": "European Journal of Biochemistry",
  "original-title": [],
  "language": "en",
  "link": [
    {
      "URL": "https://api.wiley.com/onlinelibrary/tdm/v1/articles/10.1111%2Fj.1432-1033.1973.tb02660.x",
      "content-type": "unspecified",
      "content-version": "vor",
      "intended-application": "text-mining"
    },
    {
      "URL": "https://febs.onlinelibrary.wiley.com/doi/pdf/10.1111/j.1432-1033.1973.tb02660.x",
      "content-type": "unspecified",
      "content-version": "vor",
      "intended-application": "similarity-checking"
    }
  ],
  "deposited": {
    "date-parts": [
      [
        2023,
        11,
        23
      ]
    ],
    "date-time": "2023-11-23T15:06:36Z",
    "timestamp": 1700751996000
  },
  "score": 1,
  "resource": {
    "primary": {
      "URL": "https://febs.onlinelibrary.wiley.com/doi/10.1111/j.1432-1033.1973.tb02660.x"
    }
  },
  "subtitle": [],
  "short-title": [],
  "issued": {
    "date-parts": [
      [
        1973,
        2
      ]
    ]
  },
  "references-count": 24,
  "journal-issue": {
    "issue": "1",
    "published-print": {
      "date-parts": [
        [
          1973,
          2
        ]
      ]
    }
  },
  "alternative-id": [
    "10.1111/j.1432-1033.1973.tb02660.x"
  ],
  "URL": "http://dx.doi.org/10.1111/j.1432-1033.1973.tb02660.x",
  "relation": {},
  "ISSN": [
    "0014-2956",
    "1432-1033"
  ],
  "subject": [],
  "container-title-short": "European Journal of Biochemistry",
  "published": {
    "date-parts": [
      [
        1973,
        2
      ]
    ]
  }
}