Abstract: The invention relates to a novel microorganism, designated Sphingomonas sp. strain AD109, which is capable of selectively desulfurizing dibenzothiophene. The invention also includes isolated proteins and nucleic acid sequences obtained from this microorganism. In another embodiment, the invention provides a method of using this microorganism or enzyme preparations derived therefrom in the biocatalytic desulfurization of a fossil fuel containing organic sulfur compounds.

Abstract: The present invention provides novel recombinant pseudomonads which contain a heterologous nucleic acid molecule comprising a nucleotide sequence encoding one or more desulfurization enzymes which are components of a biodesulfurization catalyst. The invention also provides a method of desulfurizing a carbonaceous material, such as a fossil fuel, which comprises organosulfur compounds. The method includes the steps of The method comprises the steps of (1) contacting the fossil fuel with an aqueous phase containing a recombinant biocatalyst which is capable of cleaving carbon-sulfur bonds and, optionally, a flavoprotein, thereby forming a fossil fuel and aqueous phase mixture; (2) maintaining the mixture under conditions sufficient for cleavage of the carbon-sulfur bonds of the organosulfur molecules by the biocatalyst, thereby resulting in a fossil fuel having a reduced organic sulfur content; and (3) separating the fossil fuel having a reduced organic sulfur content from the resulting aqueous phase.

Abstract: A novel system for cloning and expression of genes in gram-positive bacteria. The expression system is based on the finding that many gram-positive bacteria sort proteins to their cell surface through cis-acting N-terminal signal sequences and C-terminal anchor regions. In particular, the cell sorting signals of the streptococcal M6 protein, a well-known surface molecule, are used to construct a gram-positive expression system, designated SPEX (Streptococcal Protein Expression). Expression is achieved by cloning the gene of interest into an appropriate SPEX cassette which is then stably introduced into a bacterial host, such as the human commensal Streptococcus gordonii. Depending on the SPEX vector used, recombinant proteins can be anchored to the cell wall prior to release by specific endoproteolytic cleavage or secreted into the culture medium during bacterial growth. The use of host bacteria lacking extracellular proteases should protect secreted proteins from proteolytic degradation.