10.1021/acsomega.7b01435.s001 Ayesha Kashif Ayesha Kashif Lan-Huong Tran Lan-Huong Tran Sei-Heon Jang Sei-Heon Jang ChangWoo Lee ChangWoo Lee Roles of Active-Site Aromatic Residues in Cold Adaptation of Sphingomonas glacialis Esterase EstSP1 American Chemical Society 2017 Tyr 191 mutants stability enzyme activity Y 191W Arctic zone active-site walls Cold Adaptation wild-type EstSP 1. psychrophilic enzymes moderate-temperature esterases extremophilic esterases temperature adaptation Tyr 191 phenyl hydroxyl group Trp residue Tyr residue protein structure Sphingomonas glacialis Esterase EstSP 1 Trp residues Y 191H mutants novel insight PAMC Active-Site Aromatic Residues EstSP 1 psychrophilic esterase EstSP 1 2017-12-07 20:18:39 Journal contribution https://acs.figshare.com/articles/journal_contribution/Roles_of_Active-Site_Aromatic_Residues_in_Cold_Adaptation_of_Sphingomonas_glacialis_Esterase_EstSP1/5682634 The aromatic amino acids, Tyr or Trp, which line the active-site walls of esterases, stabilize the catalytic His loop via hydrogen bonding. A Tyr residue is preferred in extremophilic esterases (psychrophilic or hyperthermophilic esterases), whereas a Trp residue is preferred in moderate-temperature esterases. Here, we provide evidence that Tyr and Trp play distinct roles in cold adaptation of the psychrophilic esterase EstSP1 isolated from an Arctic bacterium Sphingomonas glacialis PAMC 26605. Stern–Volmer plots showed that the mutation of Tyr191 to Ala, Phe, Trp, and His resulted in reduced conformational flexibility of the overall protein structure. Interestingly, the Y191W and Y191H mutants showed increased thermal stability at moderate temperatures. All Tyr191 mutants showed reduced catalytic activity relative to wild-type EstSP1. Our results indicate that Tyr with its phenyl hydroxyl group is favored for increased conformational flexibility and high catalytic activity of EstSP1 at low temperatures at the expense of thermal stability. The results of this study suggest that, in the permanently cold Arctic zone, enzyme activity has been selected for psychrophilic enzymes over thermal stability. The results presented herein provide novel insight into the roles of Tyr and Trp residues for temperature adaptation of enzymes that function at low, moderate, and high temperatures.