Limits...
Oligomerization as a strategy for cold adaptation: Structure and dynamics of the GH1 β-glucosidase from Exiguobacterium antarcticum B7.

Zanphorlin LM, de Giuseppe PO, Honorato RV, Tonoli CC, Fattori J, Crespim E, de Oliveira PS, Ruller R, Murakami MT - Sci Rep (2016)

Bottom Line: Psychrophilic enzymes evolved from a plethora of structural scaffolds via multiple molecular pathways.We discovered that the selective pressure of low temperatures favored mutations that redesigned the protein surface, reduced the number of salt bridges, exposed more hydrophobic regions to the solvent and gave rise to a tetrameric arrangement not found in mesophilic and thermophilic homologues.The tetramer stabilizes the native conformation of the enzyme, leading to a 10-fold higher activity compared to the disassembled monomers.

View Article: PubMed Central - PubMed

Affiliation: Brazilian Bioethanol Science and Technology Laboratory, Campinas, São Paulo, Brazil.

ABSTRACT
Psychrophilic enzymes evolved from a plethora of structural scaffolds via multiple molecular pathways. Elucidating their adaptive strategies is instrumental to understand how life can thrive in cold ecosystems and to tailor enzymes for biotechnological applications at low temperatures. In this work, we used X-ray crystallography, in solution studies and molecular dynamics simulations to reveal the structural basis for cold adaptation of the GH1 β-glucosidase from Exiguobacterium antarcticum B7. We discovered that the selective pressure of low temperatures favored mutations that redesigned the protein surface, reduced the number of salt bridges, exposed more hydrophobic regions to the solvent and gave rise to a tetrameric arrangement not found in mesophilic and thermophilic homologues. As a result, some solvent-exposed regions became more flexible in the cold-adapted tetramer, likely contributing to enhance enzymatic activity at cold environments. The tetramer stabilizes the native conformation of the enzyme, leading to a 10-fold higher activity compared to the disassembled monomers. According to phylogenetic analysis, diverse adaptive strategies to cold environments emerged in the GH1 family, being tetramerization an alternative, not a rule. These findings reveal a novel strategy for enzyme cold adaptation and provide a framework for the semi-rational engineering of β-glucosidases aiming at cold industrial processes.

No MeSH data available.


Related in: MedlinePlus

Residues involved in glucose tolerance are conserved in the aglycone-binding site of EaBglA.(A) EaBglA superimposed on the PpBglA (PDB ID: 1E4I) and PpBglB-complexed with thiocellobiose–(PDB ID: 2O9R) from P. polymyxa. (B) Structural alignment between three eukaryotic β-glucosidases: the fungal HiBgl (PDB ID: 4MDP)12, the plant OsBgl (PDB ID: 2RGL)15 and the insect NkBgl (complexed with cellobiose; PDB ID: 3VIK)59. The highly conserved residue Trp323 has an orientation exclusively found in bacterial enzymes (A) that induces a different conformation of the reducing-end sugar compared to eukaryotic enzymes (B). The residues Trp164 and Leu179, suggested to play a role in glucose tolerance in HiBgl (B) and other GH1 enzymes1224, are also conserved in EaBglA (A).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4815018&req=5

f6: Residues involved in glucose tolerance are conserved in the aglycone-binding site of EaBglA.(A) EaBglA superimposed on the PpBglA (PDB ID: 1E4I) and PpBglB-complexed with thiocellobiose–(PDB ID: 2O9R) from P. polymyxa. (B) Structural alignment between three eukaryotic β-glucosidases: the fungal HiBgl (PDB ID: 4MDP)12, the plant OsBgl (PDB ID: 2RGL)15 and the insect NkBgl (complexed with cellobiose; PDB ID: 3VIK)59. The highly conserved residue Trp323 has an orientation exclusively found in bacterial enzymes (A) that induces a different conformation of the reducing-end sugar compared to eukaryotic enzymes (B). The residues Trp164 and Leu179, suggested to play a role in glucose tolerance in HiBgl (B) and other GH1 enzymes1224, are also conserved in EaBglA (A).

Mentions: Besides being a cold-active enzyme, EaBglA is also glucose tolerant14; a desirable feature in bioprocesses with high production of glucose, such as biomass saccharification23. Interestingly, the residues Trp164, Leu169 and His176, considered to play a role in the glucose tolerance of some GH1 β-glucosidases1224, are conserved in EaBglA. The amino acid triad X-Leu-His (X for aromatic residues) is also conserved in the more glucose-tolerant BglB from Paenibacillus polymyxa25, while the less glucose-tolerant BglA from the same bacterium26 has a valine replacing His176, supporting the influence of this residue in glucose tolerance (Fig. 6A).


Oligomerization as a strategy for cold adaptation: Structure and dynamics of the GH1 β-glucosidase from Exiguobacterium antarcticum B7.

Zanphorlin LM, de Giuseppe PO, Honorato RV, Tonoli CC, Fattori J, Crespim E, de Oliveira PS, Ruller R, Murakami MT - Sci Rep (2016)

Residues involved in glucose tolerance are conserved in the aglycone-binding site of EaBglA.(A) EaBglA superimposed on the PpBglA (PDB ID: 1E4I) and PpBglB-complexed with thiocellobiose–(PDB ID: 2O9R) from P. polymyxa. (B) Structural alignment between three eukaryotic β-glucosidases: the fungal HiBgl (PDB ID: 4MDP)12, the plant OsBgl (PDB ID: 2RGL)15 and the insect NkBgl (complexed with cellobiose; PDB ID: 3VIK)59. The highly conserved residue Trp323 has an orientation exclusively found in bacterial enzymes (A) that induces a different conformation of the reducing-end sugar compared to eukaryotic enzymes (B). The residues Trp164 and Leu179, suggested to play a role in glucose tolerance in HiBgl (B) and other GH1 enzymes1224, are also conserved in EaBglA (A).
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4815018&req=5

f6: Residues involved in glucose tolerance are conserved in the aglycone-binding site of EaBglA.(A) EaBglA superimposed on the PpBglA (PDB ID: 1E4I) and PpBglB-complexed with thiocellobiose–(PDB ID: 2O9R) from P. polymyxa. (B) Structural alignment between three eukaryotic β-glucosidases: the fungal HiBgl (PDB ID: 4MDP)12, the plant OsBgl (PDB ID: 2RGL)15 and the insect NkBgl (complexed with cellobiose; PDB ID: 3VIK)59. The highly conserved residue Trp323 has an orientation exclusively found in bacterial enzymes (A) that induces a different conformation of the reducing-end sugar compared to eukaryotic enzymes (B). The residues Trp164 and Leu179, suggested to play a role in glucose tolerance in HiBgl (B) and other GH1 enzymes1224, are also conserved in EaBglA (A).
Mentions: Besides being a cold-active enzyme, EaBglA is also glucose tolerant14; a desirable feature in bioprocesses with high production of glucose, such as biomass saccharification23. Interestingly, the residues Trp164, Leu169 and His176, considered to play a role in the glucose tolerance of some GH1 β-glucosidases1224, are conserved in EaBglA. The amino acid triad X-Leu-His (X for aromatic residues) is also conserved in the more glucose-tolerant BglB from Paenibacillus polymyxa25, while the less glucose-tolerant BglA from the same bacterium26 has a valine replacing His176, supporting the influence of this residue in glucose tolerance (Fig. 6A).

Bottom Line: Psychrophilic enzymes evolved from a plethora of structural scaffolds via multiple molecular pathways.We discovered that the selective pressure of low temperatures favored mutations that redesigned the protein surface, reduced the number of salt bridges, exposed more hydrophobic regions to the solvent and gave rise to a tetrameric arrangement not found in mesophilic and thermophilic homologues.The tetramer stabilizes the native conformation of the enzyme, leading to a 10-fold higher activity compared to the disassembled monomers.

View Article: PubMed Central - PubMed

Affiliation: Brazilian Bioethanol Science and Technology Laboratory, Campinas, São Paulo, Brazil.

ABSTRACT
Psychrophilic enzymes evolved from a plethora of structural scaffolds via multiple molecular pathways. Elucidating their adaptive strategies is instrumental to understand how life can thrive in cold ecosystems and to tailor enzymes for biotechnological applications at low temperatures. In this work, we used X-ray crystallography, in solution studies and molecular dynamics simulations to reveal the structural basis for cold adaptation of the GH1 β-glucosidase from Exiguobacterium antarcticum B7. We discovered that the selective pressure of low temperatures favored mutations that redesigned the protein surface, reduced the number of salt bridges, exposed more hydrophobic regions to the solvent and gave rise to a tetrameric arrangement not found in mesophilic and thermophilic homologues. As a result, some solvent-exposed regions became more flexible in the cold-adapted tetramer, likely contributing to enhance enzymatic activity at cold environments. The tetramer stabilizes the native conformation of the enzyme, leading to a 10-fold higher activity compared to the disassembled monomers. According to phylogenetic analysis, diverse adaptive strategies to cold environments emerged in the GH1 family, being tetramerization an alternative, not a rule. These findings reveal a novel strategy for enzyme cold adaptation and provide a framework for the semi-rational engineering of β-glucosidases aiming at cold industrial processes.

No MeSH data available.


Related in: MedlinePlus