Molecular cloning of maltooligosyltrehalose trehalohydrolase gene from Nostoc flagelliforme and trehalose-related response to stresses.

Department of Biology, College of Life and Environmental Science, Shanghai Normal University, Guilin Road 100, Shanghai,200234, People’s Republic of ChinaReceived: January 13, 2011 / Revised: May 6, 2011 / Accepted: May 25, 2011A genomic DNA fragment encoding a putativemaltooligosyltrehalose trehalohydrolase (NfMTH) fortrehalose biosynthesis was cloned by the degenerate primer-PCR from cyanobacterium Nostoc flagelliforme . The ORFof NfMTH is 1,848 bp in length and encodes 615 aminoacid residues, constituting a 70 kDa protein. The deducedamino acid sequence of NfMTH contains 4 regions highlyconserved for MTHs. By expression of NfMTH in E. coli,the function of this protein was demonstrated, where therecombinant protein catalyzed the hydrolysis of maltooligosyltrehalose to trehalose. The expressions of MTH andmaltooligosyltrehalose synthase in the filaments of N.flagelliforme were upregulated significantly under dehydrationstress, NaCl stress, and high temperature-drought stress.The accumulations of both trehalose and sucrose in thefilaments of N. flagelliforme were also improved significantlyunder the above stresses. Furthermore, trehalose accumulatedin smaller quantities than sucrose did when under NaClstress, but accumulated in higher quantities than sucrosedid when under temperature-drought stress, indicatingthat both trehalose and sucrose were involved in N.flagelliforme adapted to stresses and different strategiesconducted in response to various stress conditions.Keywords: Nostoc flagelliforme, maltooligosyltrehalosetrehalohydrolase, degenerate primer, desiccation stress, NaClstress, temperature stressNostoc flagelliforme is a kind of terrestrial and macroscopiccyanobacterium distributed in the northern and west-northern parts of Asia, Europe, and USA [5, 8]. It containshigh contents of amino acids, and anti-aging and antitumorcompounds, and has been used as a food delicacy as wellas an herbal medicine ingredient by the Chinese for centuries[8]. In nature, N. flagelliforme showed marked desiccationtolerance and extreme heat resistance. It usually grows ondry desert steppes, bare lands, or hills in dry or semi-dryareas where it often experiences severe environmentchanges such as severe drought, high temperature, andpoor and alkaline soil, under which other terrestrial Nostocspecies hardly survive [8]. Its filaments dried for a coupleof years could recover metabolic activity within hours afterrehydration [5, 9]. Its photosynthetic carbon fixation wasactive even under extremely dry conditions and strongsolar radiation in July when the land surface temperaturewas often above 44