Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: 2/16/2007
Publication Date: N/A
Citation: Interpretive Summary:
Technical Abstract: Hydrolysis of starch in cold environments by psychrophilic species of bacteria is believed to be accomplished through the production of special cold-adapted amylases. These amylases are reportedly thermally labile with low (<40 deg C) temperature optima and high specific activities at 0 to 25 deg C. Thermal lability is believed to be a consequence of enzyme adaptation to the cold, resulting from increased flexibility at the active centers, but it could also be caused by evolutionary drift in the absence of selection pressures for thermal stability. Amylase from a single psychrophilic species (Pseudoalteromonas haloplanktis) has been extensively studied. Amylases produced by two other psychrophilic species, Polaribacter glomeratus (Antarctic lake) and Gelidibacter algens (Antarctic sea ice), were examined to see if they exhibited properties similar to Pseudoalteromonas sp. and if they were suitable for low-temperature applications that were contemplated. Amylase activity was assayed using the Infinity Amylase Assay reagent [ethylidine-p-nitrophenyl maltoheptaoside (EtG7PNP, Thermo Electron, Melbourne, Australia) or dinitrosalicylic acid (starch assay). SDS-PAGE used the Laemmli procedure and precast polyacrylamide gels from BioRad, Inc (Hercules, CA). High levels of amylase activity could be released by sonicating cell suspensions. Polaribacter and Gelidibacter sp. produced amylases with molecular masses of 120 and 78 kDa, respectively, determined by SDS-PAGE. Their amylases were thermally stable (70% activity after 1 hr at 50 deg C). Temperature optima (EtG7PNP), pH optima (starch), and Km values (EtG7PNP, 20 deg C) were: 50 deg C, 6-8, 67 micromolar (Polaribacter); and 45 deg C, 6.0, 186 micromolar (Gelidibacter). Both amylases required Ca+2 for activity, but neither amylase possessed a raw starch-binding domain. Conclusions: (1) evolutionary pressures select for exoenzymes adequate for the survival needs of cold-adapted bacteria but not necessarily for labile exoenzymes with low temperature optima, (2) these amylases might be cold-adapted in spite of high temperature optima and thermal stability.