Abstract
The haloarchaeon Halorubrum lacusprofundi is among the few polyextremophilic organisms capable of surviving in one of the most extreme aquatic environments on Earth, the Deep Lake of Antarctica (−18 °C to +11.5 °C and 21–28%, w/v salt content). Hence, H. lacusprofundi has been proposed as a model for biotechnology and astrobiology to investigate potential life beyond Earth. To understand the mechanisms that allow proteins to adapt to both salinity and cold, we structurally (including X-ray crystallography and molecular dynamics simulations) and functionally characterized the β-galactosidase from H. lacusprofundi (hla_bga). Recombinant hla_bga (produced in Haloferax volcanii) revealed exceptional stability, tolerating up to 4 M NaCl and up to 20% (v/v) of organic solvents. Despite being cold-adapted, hla_bga was also stable up to 60 °C. Structural analysis showed that hla_bga combined increased surface acidity (associated with halophily) with increased structural flexibility, fine-tuned on a residue level, for sustaining activity at low temperatures. The resulting blend enhanced structural flexibility at low temperatures but also limited protein movements at higher temperatures relative to mesophilic homologs. Collectively, these observations help in understanding the molecular basis of a dual psychrophilic and halophilic adaptation and suggest that such enzymes may be intrinsically stable and functional over an exceptionally large temperature range.
Highlights
Enzymes have emerged as preferred tools in green chemistry and have the potential to play a vital role in sustainable development in chemical, biotechnological, bioremediation, agricultural and pharmaceutical industries [1,2]
Improvements were achieved by protein engineering, these procedures are often lengthy and expensive with non-generalizable outcomes, because increased enzyme stability mostly results from specific mutations, which usually do not obey any obvious trends or patterns [6,7,8,9,10,11]
Since the metabolic processes and physiological functions of extremophiles are adapted to prevail under harsh conditions, enzymes from these microorganisms, called extremozymes, possess unique features enabling them to carry out reactions under extreme conditions, such as the presence of up to 5.2 M salt, various surfactants, organic solvents, elevated or low temperature, and at alkaline pH [14,19]
Summary
Enzymes have emerged as preferred tools in green chemistry and have the potential to play a vital role in sustainable development in chemical, biotechnological, bioremediation, agricultural and pharmaceutical industries [1,2]. The high stability of enzymes towards salt entails tolerance to low water activity, such as prevailing in mixtures of aqueous and organic or non-aqueous media and low temperatures can save energy and eliminate microbial contamination [14]. These characteristics increase the enzymes’ potential as industrial biocatalysts because organic solvents are often used to improve the solubility of hydrophobic substrates, alter the hydrolytic as well as the kinetic equilibrium and have the potential to increase the yield and specificity of the product [15]. Our study provides rationales for the biological and industrial enzyme adaptation to polyextreme conditions
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