Physiological and biochemical characterization of the two α-l-rhamnosidases of Lactobacillus plantarum NCC245

Abstract

This work is believed to be the first report on the physiological and biochemical characterization of a-L-rhamnosidases in lactic acid bacteria. A total of 216 strains representing 37 species and eight genera of food-grade bacteria were screened for a-L-rhamnosidase activity. The majority of positive bacteria (25 out of 35) were Lactobacillus plantarum strains, and activity of the L. plantarum strain NCC245 was examined in more detail. The analysis of a-L-rhamnosidase activity under different growth conditions revealed dual regulation of the enzyme activity, involving carbon catabolite repression and induction: the enzyme activity was downregulated by glucose and upregulated by L-rhamnose. The expression of the two a-L-rhamnosidase genes rhaB1 and rhaB2 and two predicted permease genes rhaP1 and rhaP2, identified in a probable operon rhaP2B2P1B1, was repressed by glucose and induced by L-rhamnose, showing regulation at the transcriptional level. The two a-L-rhamnosidase genes were overexpressed and purified from Escherichia coli. RhaB1 activity was maximal at 50 6C and at neutral pH and RhaB2 maximal activity was detected at 60 6C and at pH 5, with high residual activity at 70 6C. Both enzymes showed a preference for the a-1,6 linkage of L-rhamnose to b-D-glucose, hesperidin and rutin being their best substrates, but, surprisingly, no activity was detected towards the a-1,2 linkage in naringin under the tested conditions. In conclusion, we identified and characterized the strain L. plantarum NCC245 and its two a-L-rhamnosidase enzymes, which might be applied for improvement of bioavailability of health-beneficial polyphenols, such as hesperidin, in humans.

The Wood Rot Ascomycete Xylaria polymorpha Produces a Novel GH78 Glycoside Hydrolase That Exhibits α- l -Rhamnosidase and Feruloyl Esterase Activities and Releases Hydroxycinnamic Acids from Lignocelluloses

“…The native protein may be a heterodimer that consists of two subunits (39 and 62 kDa), or it is rather labile and may partially split under denaturing conditions. Similar observations were made for different recombinant bacterial rhamnosidases which can occur as monomers and dimers depending on native or denaturing conditions (3,8,50).…”

Characterization of Two Distinct Glycosyl Hydrolase Family 78 α- l -Rhamnosidases from Pediococcus acidilactici

“…However, only a few bacterial rhamnosidases have been characterized (15,19,20,54) to date, and the properties of rhamnosidases derived from food LAB (Lactobacillus plantarum and Lactobacillus acidophilus) have only recently been reported (3,7). In the present study, we characterized two ␣-L-rhamnosidases derived from the homofermentative lactic acid bacterium Pediococcus acidilactici.…”

Evaluation and characterization of new α-L-rhamnosidase-producing yeast strains

“…The optimum temperature obtained was similar to the α-L-rhamnosidases from Bacillus sp. GL1 (Hashimoto et al, 1999), Lactobacillus plantarum NCC 245 (Avila et al, 2009), Aspergillus flavus (Scaroni et al, 2002), Curvularia lunata (Feng et al, 2007), Aspergillus niger (Puri and Kalra, 2005) and Aspergillus kawachii (Koseki et al, 2008). Enzymes from Pichia angusta (Yanai and Sato, 2000), Aspergillus kawachii, Penicillium aureatiogriseum and Trichoderma longibrachiatu (Scaroni et al, 2002) were optimally active at 40°C and 60°C.…”