Residue Asn277 Affects the Stability and Substrate Specificity of the SMG1 Lipase from Malassezia globosa

Lan, Dongming; Wang, Qian; Xu, Jinxin; Zhou, Pengfei; Yang, Bo; Wang, Yonghua

doi:10.3390/ijms16047273

Cited by 16 publications

(9 citation statements)

References 36 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…A, variant F278N retains the same overall structural architecture as the wild type, except for some minor small angle dislocation in strands β1, the preceding loop and β4. Residues 277 and 288 that were previously reported from our group to contribute to the stability and substrate specificity are in close vicinity of residue 278 and do not show any structural changes. The same is observed for the lid region (residues 101–119), which superimposes completely with that in the closed form (PDB ID: http://www.rcsb.org/pdb/search/structidSearch.do?structureId=3UUE) and displays relatively low B ‐factor (Fig.…”

Section: Resultssupporting

confidence: 58%

Structure of product‐bound SMG1 lipase: active site gating implications

Guo

Pavlidis

et al. 2015

The FEBS Journal

Self Cite

View full text Add to dashboard Cite

Monoacylglycerol and diacylglycerol lipases are industrially interesting enzymes, due to the health benefits that arise from the consumption of diglycerides compared to the traditional triglyceride oils. Most lipases possess an a-helix (lid) directly over the catalytic pocket which regulates the activity of the enzyme. Generally, lipases exist in active and inactive conformations, depending on the positioning of this lid subdomain. However, lipase SMG1, a monoacylglycerol and diacylglycerol specific lipase, has an atypical activation mechanism. In the present study we were able to prove by crystallography, in silico analysis and activity tests that only two positions, residues 102 and 278, are responsible for a gating mechanism that regulates the active and inactive states of the lipase, and that no significant structural changes take place during activation except for oxyanion hole formation. The elucidation of the gating effect provided data enabling the rational design of improved lipases with 6-fold increase in the hydrolytic activity toward diacylglycerols, just by providing additional substrate stabilization with a single mutation (F278N or F278T). Due to the conservation of F278 among the monoacylglycerol and diacylglycerol lipases in the Rhizomucor miehei lipase-like family, the gating mechanism described herein might represent a general mechanism applicable to other monoacylglycerol and diacylglycerol lipases as well.

show abstract

Section: Resultssupporting

confidence: 58%

Structure of product‐bound SMG1 lipase: active site gating implications

Guo

Pavlidis

et al. 2015

The FEBS Journal

Self Cite

View full text Add to dashboard Cite

show abstract

“…Currently, many protein engineering methods have been developed to improve esterases (or lipases) properties, including substrate specificity [41,42], activity [43,44], thermostability [45], and enantioselectivity [46,47]. However, limited studies for enhancing alkali tolerance have been reported, especially based on structural information.…”

Section: Discussionmentioning

confidence: 99%

Structure-guided protein engineering increases enzymatic activities of the SGNH family esterases

Huo

et al. 2020

Biotechnol Biofuels

View full text Add to dashboard Cite

Background: Esterases and lipases hydrolyze short-chain esters and long-chain triglycerides, respectively, and therefore play essential roles in the synthesis and decomposition of ester bonds in the pharmaceutical and food industries. Many SGNH family esterases share high similarity in sequences. However, they have distinct enzymatic activities toward the same substrates. Due to a lack of structural information, the detailed catalytic mechanisms of these esterases remain barely investigated. Results: In this study, we identified two SGNH family esterases, CrmE10 and AlinE4, from marine bacteria with significantly different preferences for pH, temperature, metal ion, and organic solvent tolerance despite high sequence similarity. The crystal structures of these two esterases, including wild type and mutants, were determined to high resolutions ranging from 1.18 Å to 2.24 Å. Both CrmE10 and AlinE4 were composed of five β-strands and nine α-helices, which formed one compact N-terminal α/β globular domain and one extended C-terminal domain. The aspartic residues (D178 in CrmE10/D162 in AlinE4) destabilized the conformations of the catalytic triad (Ser-Asp-His) in both esterases, and the metal ion Cd 2+ might reduce enzymatic activity by blocking proton transfer or substrate binding. CrmE10 and AlinE4 showed distinctly different electrostatic surface potentials, despite the similar atomic architectures and a similar swap catalytic mechanism. When five negatively charged residues (Asp or Glu) were mutated to residue Lys, CrmE10 obtained elevated alkaline adaptability and significantly increased the enzymatic activity from 0 to 20% at pH 10.5. Also, CrmE10 mutants exhibited dramatic change for enzymatic properties when compared with the wide-type enzyme. Conclusions: These findings offer a perspective for understanding the catalytic mechanism of different esterases and might facilitate the industrial biocatalytic applications.

show abstract

“…Currently, many protein engineering methods have been developed to improve esterases (or lipases) properties, including substrate speci city [41,42], activity [43,44], thermostability [45], and enantioselectivity [46,47]. However, limited studies for enhancing alkali-tolerance have been reported, especially based on structural information.…”

Section: Discussionmentioning

confidence: 99%

Structure guided protein engineering increases enzymatic activities of the SGNH family esterases

Huo

et al. 2020

Preprint

View full text Add to dashboard Cite

Background Esterases and lipases hydrolyze short-chain esters and long-chain triglycerides, respectively, and therefore play essential roles in the synthesis and decomposition of ester bonds in the pharmaceutical and food industries. Many SGNH family esterases share high similarity in sequences. However, they have distinct enzymatic activities toward the same substrates. Due to a lack of structural information, the detailed catalytic mechanisms of these esterases remain barely investigated. Results In this study, we identified two SGNH family esterases, CrmE10 and AlinE4, from marine bacteria with significantly different preferences for pH, temperature, metal ion, and organic solvent tolerance despite high sequence similarity. The crystal structures of these two esterases, including wild type and mutants, were determined to high resolutions ranging from 1.18 Å to 2.24 Å. Both CrmE10 and AlinE4 were composed of five β-strands and nine α-helices, which formed one compact N-terminal α/β globular domain and one extended C-terminal domain. The aspartic residues (D178 in CrmE10/ D162 in AlinE4) destabilized the conformations of the catalytic triad (Ser-Asp-His) in both esterases, and the metal ion Cd2+ might reduce enzymatic activity by blocking proton transfer or substrate binding. CrmE10 and AlinE4 showed distinctly different electrostatic surface potentials, despite the similar atomic architectures and a similar swap catalytic mechanism. When five negative charged residues (Asp or Glu) were mutated to residue Lys, CrmE10 obtained elevated alkaline-adaptability and significantly increased the enzymatic activity from 0% to 20% at pH 10.5. Also, CrmE10 mutants exhibited dramatic changes for enzymatic properties when compared with the wide-type enzyme. Conclusions These findings offer a perspective for understanding the catalytic mechanism of different esterases and might facilitate the industrial biocatalytic applications.

show abstract

Residue Asn277 Affects the Stability and Substrate Specificity of the SMG1 Lipase from Malassezia globosa

Cited by 16 publications

References 36 publications

Structure of product‐bound SMG1 lipase: active site gating implications

Structure of product‐bound SMG1 lipase: active site gating implications

Structure-guided protein engineering increases enzymatic activities of the SGNH family esterases

Structure guided protein engineering increases enzymatic activities of the SGNH family esterases

Contact Info

Product

Resources

About