Psychrobacter 273-4 was isolated from a 20,000-40,000-year-old Siberian permafrost core, which is characterized by low temperature, low water activity, and high salinity. To explore how 273-4 survives in the permafrost environment, proteins in four 273-4 samples cultured at 4 and 22 degrees C in media with and without 5% sodium chloride were profiled and comparatively studied using 2-D HPLC and MS. The method used herein involved fractionation via a pH gradient using chromatofocusing followed by nonporous silica (NPS) RP-HPLC and on-line electrospray mass mapping. It was observed that 33 proteins were involved in the adaptation to low temperature in the cells grown in the nonsaline media while there were only 14 proteins involved in the saline media. There were 45 proteins observed differentially expressed in response to salt at 22 degrees C while there were 22 proteins at 4 degrees C. In addition, 5% NaCl and 4 degrees C showed a combination effect on protein expression. A total of 56 proteins involved in the adaptation to low temperature and salt were identified using MS and database searching. The differentially expressed proteins were classified into different functional categories where the response of the regulation system to stress appears to be very elaborate. The evidence shows that the adaptation of 273-4 is based primarily on the control of translation and transcription, the synthesis of proteins (chaperones) to facilitate RNA and protein folding, and the regulation of metabolic pathways.