CLONING AND SEQUENCING OF THE SPHINGOMONAS (PSEUDOMONAS) PAUCIMOBILISGENE ESSENTIAL FOR THE O-DEMETHYLATION OF VANILLATE AND SYRINGATE

Sphingomonas (Pseudomonas) paucimobilis SYK-6 is able to grow on 5,5'- dehydrodivanillic acid (DDVA), syringate, vanillate, and other dimeric model compounds of lignin as a sole carbon source, Nitrosoguanidine m utagenesis of S. paucimobilis SYK-6 was performed, and two mutants wit h altered DDVA degradation pathways were isolated. The mutant strain N T-1 could not degrade DDVA, but could degrade syringate, vanillate, an d ,2',3'-trihydroxy-3-methoxy-5,5'-dicarboxybiphenyl (OH-DDVA), Strain DC-49 could slowly assimilate DDVA, but could degrade neither vanilla te nor syringate, although it could degrade protocatechuate and 3-O-me thylgallate. A complementing DNA fragment of strain DC-49 was isolated from the cosmid library of strain SYK-6. The minimum DNA fragment com plementing DC-49 was determined to be the 1.8-kbp insert of pKEX2.0, S equencing analysis showed an open reading frame of 1,671 bp in this fr agment, and a similarity search indicated that the deduced amino acid sequence of this open reading frame had significant similarity (60%) t o the formyltetrahydrofolate synthetase of Clostridium thermoaceticum.