Heterologous production of (-)-geosmin in Saccharomyces cerevisiae.

(-)-Geosmin has high demand in perfumes and cosmetic products for its earthy congenial aroma. The current production of (-)-geosmin is either by distillation of sun-baked soil or by inefficient chemical synthesis because of the presence of multiple chiral centers. Fermentation processes are not viable as the titers of the Streptomyces sp. based processes are low. This work presents an alternative route by the heterologous synthesis of (-)-geosmin in Saccharomyces cerevisiae. The enzyme involved is the bifunctional geosmin synthase that catalyzes the conversion of farnesyl diphosphate to germacradienol and germacradienol to geosmin. This study evaluated the activity of many orthologs of geosmin synthase when expressed heterologously in S. cerevisiae. When the well-characterized CAB41566 from Streptomyces coelicolor origin was tested, germacradienol and germacrene D were detected but no geosmin. Bioinformatic analysis based on high/low identities to N-terminal and C-terminal domains of CAB41566 was carried out to identify different orthologs of geosmin synthase proteins from different bacterial and fungal origins. ADO68918 of Stigmatella aurantiaca origin showed the best activity among the tested orthologs, not only in terms of geosmin production but also an order of magnitude higher total abundance of the products of geosmin synthase as compared to CAB41566. This study successfully demonstrated the production of (-)-geosmin in S. cerevisiae and offers an alternative, sustainable and environment-friendly approach to producing (-)-geosmin.

Selection of methanotrophic platform for methanol production using methane and biogas.

To develop biotechnological process for methane to methanol conversion, selection of a suitable methanotrophic platform is an important aspect. Systematic approach based on literature and public databases was developed to select representative methanotrophs Methylotuvimicrobium alcaliphilum, Methylomonas methanica, Methylosinus trichosporium and Methylocella silvestris. Selected methanotrophs were further investigated for methanol tolerance and methanol production on pure methane as well as biogas along with key enzyme activities involved in methane utilization. Among selected methanotrophs M. alcaliphilum showed maximum methanol tolerance of 6% v/v along with maximum methanol production of 307.90 mg/L and 247.37 mg/L on pure methane and biogas respectively. Activity of methane monooxygenase and formate dehydrogenase enzymes in M.alcaliphilum was significantly higher up to 98.40 nmol/min/mg cells and 0.87 U/mg protein, respectively. Biotransformation trials in 14 L fermentor resulted in increased methanol production up to 418 and 331.20 mg/L, with yield coefficient 0.83 and 0.71 mg methanol/mg of pure methane and biogas respectively. The systematic selection resulted in haloalkaliphilic strain M. alcaliphilum as one of the potential methanotroph for bio-methanol production.