On the effect of mutations in bovine or camel chymosin on the thermodynamics of binding κ-caseins

Samiul M. Ansari, Jesper Sørensen, Birgit Schiøtt, David S. Palmer

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Abstract

Bovine and camel chymosins are aspartic proteases that are used in dairy food manufacturing. Both enzymes catalyse proteolysis of a milk protein, κ-casein, which helps to initiate milk coagulation. Surprisingly, camel chymosin shows a 70 % higher clotting activity than bovine chymosin for bovine milk, while exhibiting only 20 % of the unspecific proteolytic activity. By contrast, bovine chymosin is a poor coagulant for camel milk. Although both enzymes are marketed commercially, the disparity in their catalytic activity is not yet well understood at a molecular level, due in part to a lack of atomistic resolution data about the chymosin - κ-casein complexes. Here, we report computational alanine scanning calculations of all four chymosin - κ-casein complexes, allowing us to elucidate the influence that individual residues have on binding thermodynamics. Of the 12 sequence di erences in the binding sites of bovine and camel chymosin, eight are shown to be particularly important for understanding di erences in the binding thermodynamics (Asp112Glu, Lys221Val, Gln242Arg, Gln278Lys. Glu290Asp, His292Asn, Gln294Glu, and Lys295Leu. Residue in bovine chymosin written first). The relative binding free energies of single-point mutants of chymosin are calculated using the molecular mechanics three dimensional reference interaction site model (MM-3DRISM). Visualisation of the solvent density functions calculated by 3DRISM reveals the di erence in solvation of the binding sites of chymosin mutants.
Original languageEnglish
Pages (from-to)1-36
Number of pages36
JournalProteins: Structure, Function, and Bioinformatics
Early online date30 Oct 2017
DOIs
Publication statusE-pub ahead of print - 30 Oct 2017

Fingerprint

Chymosin
Camelus
Caseins
Thermodynamics
Mutation
Milk
Binding Sites
Proteolysis
Coagulants
Molecular mechanics
Dairies
Milk Proteins
Solvation
Enzymes
Coagulation
Mechanics
Alanine
Probability density function
Free energy
Catalyst activity

Keywords

  • chymosin
  • cheese
  • aspartic protease
  • molecular simulation
  • MM3DRISM
  • free energy
  • alanine scanning
  • k-casein

Cite this

Ansari, Samiul M. ; Sørensen, Jesper ; Schiøtt, Birgit ; Palmer, David S. / On the effect of mutations in bovine or camel chymosin on the thermodynamics of binding κ-caseins. In: Proteins: Structure, Function, and Bioinformatics. 2017 ; pp. 1-36.
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title = "On the effect of mutations in bovine or camel chymosin on the thermodynamics of binding κ-caseins",
abstract = "Bovine and camel chymosins are aspartic proteases that are used in dairy food manufacturing. Both enzymes catalyse proteolysis of a milk protein, κ-casein, which helps to initiate milk coagulation. Surprisingly, camel chymosin shows a 70 {\%} higher clotting activity than bovine chymosin for bovine milk, while exhibiting only 20 {\%} of the unspecific proteolytic activity. By contrast, bovine chymosin is a poor coagulant for camel milk. Although both enzymes are marketed commercially, the disparity in their catalytic activity is not yet well understood at a molecular level, due in part to a lack of atomistic resolution data about the chymosin - κ-casein complexes. Here, we report computational alanine scanning calculations of all four chymosin - κ-casein complexes, allowing us to elucidate the influence that individual residues have on binding thermodynamics. Of the 12 sequence di erences in the binding sites of bovine and camel chymosin, eight are shown to be particularly important for understanding di erences in the binding thermodynamics (Asp112Glu, Lys221Val, Gln242Arg, Gln278Lys. Glu290Asp, His292Asn, Gln294Glu, and Lys295Leu. Residue in bovine chymosin written first). The relative binding free energies of single-point mutants of chymosin are calculated using the molecular mechanics three dimensional reference interaction site model (MM-3DRISM). Visualisation of the solvent density functions calculated by 3DRISM reveals the di erence in solvation of the binding sites of chymosin mutants.",
keywords = "chymosin, cheese, aspartic protease, molecular simulation, MM3DRISM, free energy, alanine scanning, k-casein",
author = "Ansari, {Samiul M.} and Jesper S{\o}rensen and Birgit Schi{\o}tt and Palmer, {David S.}",
note = "This is the peer reviewed version of the following article: Ansari, SM, S{\o}rensen, J, Schi{\o}tt, B & Palmer, DS 2017, 'On the effect of mutations in bovine or camel chymosin on the thermodynamics of binding κ-caseins' Proteins: Structure, Function, and Bioinformatics, pp. 1-36., which has been published in final form at https://doi.org/10.1002/prot.25410. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.",
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Ansari, SM, Sørensen, J, Schiøtt, B & Palmer, DS 2017, 'On the effect of mutations in bovine or camel chymosin on the thermodynamics of binding κ-caseins', Proteins: Structure, Function, and Bioinformatics, pp. 1-36. https://doi.org/10.1002/prot.25410

On the effect of mutations in bovine or camel chymosin on the thermodynamics of binding κ-caseins. / Ansari, Samiul M.; Sørensen, Jesper; Schiøtt, Birgit; Palmer, David S.

In: Proteins: Structure, Function, and Bioinformatics, 30.10.2017, p. 1-36.

Research output: Contribution to journal › Article

TY - JOUR

T1 - On the effect of mutations in bovine or camel chymosin on the thermodynamics of binding κ-caseins

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AU - Sørensen, Jesper

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AU - Palmer, David S.

N1 - This is the peer reviewed version of the following article: Ansari, SM, Sørensen, J, Schiøtt, B & Palmer, DS 2017, 'On the effect of mutations in bovine or camel chymosin on the thermodynamics of binding κ-caseins' Proteins: Structure, Function, and Bioinformatics, pp. 1-36., which has been published in final form at https://doi.org/10.1002/prot.25410. This article may be used for non-commercial purposes in accordance with Wiley Terms and Conditions for Self-Archiving.

PY - 2017/10/30

Y1 - 2017/10/30

N2 - Bovine and camel chymosins are aspartic proteases that are used in dairy food manufacturing. Both enzymes catalyse proteolysis of a milk protein, κ-casein, which helps to initiate milk coagulation. Surprisingly, camel chymosin shows a 70 % higher clotting activity than bovine chymosin for bovine milk, while exhibiting only 20 % of the unspecific proteolytic activity. By contrast, bovine chymosin is a poor coagulant for camel milk. Although both enzymes are marketed commercially, the disparity in their catalytic activity is not yet well understood at a molecular level, due in part to a lack of atomistic resolution data about the chymosin - κ-casein complexes. Here, we report computational alanine scanning calculations of all four chymosin - κ-casein complexes, allowing us to elucidate the influence that individual residues have on binding thermodynamics. Of the 12 sequence di erences in the binding sites of bovine and camel chymosin, eight are shown to be particularly important for understanding di erences in the binding thermodynamics (Asp112Glu, Lys221Val, Gln242Arg, Gln278Lys. Glu290Asp, His292Asn, Gln294Glu, and Lys295Leu. Residue in bovine chymosin written first). The relative binding free energies of single-point mutants of chymosin are calculated using the molecular mechanics three dimensional reference interaction site model (MM-3DRISM). Visualisation of the solvent density functions calculated by 3DRISM reveals the di erence in solvation of the binding sites of chymosin mutants.

AB - Bovine and camel chymosins are aspartic proteases that are used in dairy food manufacturing. Both enzymes catalyse proteolysis of a milk protein, κ-casein, which helps to initiate milk coagulation. Surprisingly, camel chymosin shows a 70 % higher clotting activity than bovine chymosin for bovine milk, while exhibiting only 20 % of the unspecific proteolytic activity. By contrast, bovine chymosin is a poor coagulant for camel milk. Although both enzymes are marketed commercially, the disparity in their catalytic activity is not yet well understood at a molecular level, due in part to a lack of atomistic resolution data about the chymosin - κ-casein complexes. Here, we report computational alanine scanning calculations of all four chymosin - κ-casein complexes, allowing us to elucidate the influence that individual residues have on binding thermodynamics. Of the 12 sequence di erences in the binding sites of bovine and camel chymosin, eight are shown to be particularly important for understanding di erences in the binding thermodynamics (Asp112Glu, Lys221Val, Gln242Arg, Gln278Lys. Glu290Asp, His292Asn, Gln294Glu, and Lys295Leu. Residue in bovine chymosin written first). The relative binding free energies of single-point mutants of chymosin are calculated using the molecular mechanics three dimensional reference interaction site model (MM-3DRISM). Visualisation of the solvent density functions calculated by 3DRISM reveals the di erence in solvation of the binding sites of chymosin mutants.

KW - chymosin

KW - cheese

KW - aspartic protease

KW - molecular simulation

KW - MM3DRISM

KW - free energy

KW - alanine scanning

KW - k-casein

UR - http://onlinelibrary.wiley.com/journal/10.1002/(ISSN)1097-0134

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DO - 10.1002/prot.25410

M3 - Article

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EP - 36

JO - Proteins: Structure, Function, and Bioinformatics

JF - Proteins: Structure, Function, and Bioinformatics

SN - 0887-3585

ER -

Ansari SM, Sørensen J, Schiøtt B, Palmer DS. On the effect of mutations in bovine or camel chymosin on the thermodynamics of binding κ-caseins. Proteins: Structure, Function, and Bioinformatics. 2017 Oct 30;1-36. https://doi.org/10.1002/prot.25410

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