CLONING AND EXPRESSION ANALYSIS OF BETAINE ALDEHYDE DEHYDROGENASE FROM Pseudomonas fluorescens

Abstract

Pseudomonas fluorescens are gramnegative rod shaped bacteria that improve plant health and nutrition (Rodriguez and Pfender, 1997; de Bruijn et al., 2007). Most studies describe P. fluorescens as a psychrotrophic bacterium unable to grow at temperatures greater than 32C and therefore as an avirulent bacterium to humans (Naseby et al., 2001). It has many positive effects on the plant health as it enhances the production of plant growth hormones, it boosts the suppression of pathogens (especially fungi and oomycetes), and it directs the elicitation of plant defense responses (Haas and Defago, 2005). Many micro-organisms use glycine betaine (GB) as a sole carbon, nitrogen, and energy source. GB is an efficient osmoprotectant, which is accumulated to counteract drought and high salinity environments, and to maintain a positive cell turgor, needed for cell extension growth. Betaine aldehyde dehydrogenase (BADH) catalyzes the final step of glycine betaine from choline (Chen and Murata, 2002; Hanson et al., 1985; He et al., 2004). Several reports have demonstrated the purification of BADHs from several species (Arakawa et al., 1987; Hibino et al., 2001; Ishitani et al., 1993). Overexpression of BADH may protect intracellular enzymes and organelles against the elevation of intracellular ionic strength or temperature, which results in increase of tolerance of salt and osmotic stresses in many organisms (Kempf and Bremer, 1998; Welsh, 2000; Li et al., 2006). Beside its function in the production of glycine betaine, BADH is a key enzyme for the growth of many pathogens (Velasco-Garci´a et al., 2000). The growth arrest of the pathogen can be accomplished by blocking choline degradation, abolishing synthesis of the osmoprotectant glycine betaine, and accumulating the BADH substrate, betaine aldehyde, which is highly toxic to the pathogen (Boch et al., 1996; Sage et al., 1997). In this investigation, the full length of betB gene, PfBADH, coding for BADH enzyme was isolated from Pseudomonas fluorescens, cloned in Escherichia coli and identified by studying the gene sequence and its expression analysis. Cloning the full length gene of PfBADH is a starting point for the strategic improvement of commercial crops grown under biotic and abiotic stress conditions using gene transfer techniques.