Structural features of bacterial luciferase related to temperature adaptation

2022 The Thirteenth International Multiconference   unpublished
Motivation and Aim: Luminous bacteria are widely distributed in nature, with the majority of species living in the ocean and only a few representatives described as freshwater and terrestrial [1] . Thus, they have adapted to live in various environmental conditions and at different temperatures as well. Based on optimal growth temperature, psychrophilic and mesophilic species of luminous bacteria can be distinguished. The features of bacteria adaptation to spescific temperature range can be
more » ... across all levels of organization, including enzymatic. One of the most studied enzyme of luminous bacteria is luciferase, which is responsible for bioluminescent reaction and widely applied as an element of biosensors in fundamental research, biotechnological applications, ecotoxicology, etc. [2, 3] . The aim of this study is to reveal the structural features of two luciferases from different subfamilies (Photobacterium leiognathi and Vibrio harveyi) related to temperature dependence of their function. Methods and Algorithms: The set of 47 amino acid sequences of bacterial luciferase associated with fully or partially sequenced lux-operons from NCBI database was obtained mannually. We used the set of sequences to analyze the amino acid substitutions specific to temperature adaptation with GroupSim and multi-Harmony program packages. The temperature dependence of reaction kinetics for V. harveyi and P. leiognathi luciferases was measured using stopped-flow technique with SX-20 analyzer (Applied Photophysics) in the range 5-45 °C. Additionally, we have run 100ns molecular dynamics simulations for both enzymes at 5, 10, 27, 45, 60 °C using the GROMACS 2020.4 software package. Results: Sequence analysis revealed that P. leiognathi luciferase is highly homologous to luciferases from psychrophilic strains, while V. harveyi luciferase is very similar to the thermostable luciferase from V. campbellii. According to experimental results, V. harveyi luciferase does have a wider temperature optimum shifted to higher temperatures compared to P. leiognathi luciferase. However, the total quantum yeld is lower in the reaction catalyzed by V. harveyi luciferase compared to the P. leiognathi one. Kinetics analysis indicated that P. leiognathi luciferase better stabilizes 4ahydroperoxyflavin intermediate in the temperature range of 20-35 °C, and is less effective at temperatures of 40-45 °C. Computational simulation showed that P. leiognathi structure is more dynamic, but the shape of the protein does not remarkably change with increasing temperature, which is typical for psychrophilic enzymes. V. harveyi luciferase undergoes minor overall fluctuation at studied temperatures indicating its higher rigidity, which is characteristic for thermostable enzymes. We also analyzed the flexibility of sreparate structural elements of the proteins at different temperatures. The fluctuations of a mobile loop
doi:10.18699/sbb-2022-157 fatcat:xih47cbbzffe3ksodmra4t3j2e