ARS | Publication request: Carbohydrate utilization and detection of a nucleotide hydrolase in Lactobacillus buchneri

Submitted to: Meeting Abstract
Publication Type: Abstract Only
Publication Acceptance Date: November 30, 2007
Publication Date: November 30, 2007
Citation: Liu, S., Bischoff, K.M., Price, N.P., Hughes, S.R. 2007. Carbohydrate utilization and detection of a nucleotide hydrolase in Lactobacillus buchneri [abstract]. Symposium on Biocatalysis and Biotechnology. p. 49.

Technical Abstract: Lactobacillus buchneri strains NRRL 1837, DSM 5987, and NRRL B-30929 were examined for capacity to metabolize various carbohydrates via growth and fermentation analyses. Carbon sources used for this study included D-melibiose, inosine, uridine, D-melezitose, maltotriose, N-acetyl-D-glucosamine, sucrose, mannose, cellobiose, trehalose, turanose, lactulose, lactose, and raffinose. The analyses indicated that each strain is able to metabolize a particular spectrum of carbon sources. All three strains can utilize raffinose, lactulose, D-melibiose, inosine, and uridine. None of the strains are capable of utilizing mannose and N-acetyl-D-glucosamine although a previous study using biolog PM array suggested potential metabolism of these two substrates. Whereas NRRL 1837 can metabolize the following three substrates D-melezitose (trisaccharide), sucrose and turanose (disaccharide), DSM 5987 metabolizes D-melezitose and sucrose, while NRRL B-30929 metabolizes only turanose. Furthermore, NRRL 1837 and DSM 5987 are unable to metabolize cellobiose while NRRL B-30929 can metabolize cellobiose after prolonged incubation. These results allowed us to establish a unique phenotype pertaining to each strain. The capacity for using cellobiose distinguishes B-30929 from others as a novel biocatalyst for potential applications of cellulosic biomass conversion. For the substrates that support growth, HPLC analyses were also performed. Common products are lactate, ethanol, and acetate. However, an unknown compound was produced when uridine was supplied as the sole substrate in all three strains. The compound was readily precipitated from culture broth as a white pellet at refrigerator temperatures or at room temperature with higher concentrations. Analysis of this compound by GC-MS, MALDI-TOF MS, and 1H-NMR indicated that it is free uracil. Since uridine is a nucleoside that is formed when uracil is attached to a ribose ring via a beta-glycosidic bond, this result suggests that a nucleotide hydrolase is present and the enzyme is likely induced by uridine (see below). Quantitative analyses showed that the molar ratio of uracil produced to uridine used is 1.95:1. This indicated that more uracil is produced after the complete hydrolysis of the uridine supplied in media. The additional uracil could come from the hydrolysis of UMP from RNA or deoxy-thymidine from DNA. Uracil in combination with 5-fluorouacil analogs are known as effective chemotherapy drugs for several types of cancer. Fermentative production of uracil and its derivatives from cheaper biomass feedstocks will have environmental friendly advantages over the chemical synthesis currently being used in pharmarceutical industry.