Abstract
Syntrophic acetate-oxidizing bacteria have been identified as key organisms for efficient biogas production from protein-rich materials. They normally grow as lithotrophs or heterotrophs, producing acetate through the Wood-Ljungdahl pathway, but when growing in syntrophy with methanogens, they reportedly reverse this pathway and oxidize acetate to hydrogen and carbon dioxide. However, the biochemical and regulatory mechanisms behind the shift and the way in which the bacteria regain energy remain unknown. In a genome-walking approach, starting with degenerated primers, we identified those gene clusters in Syntrophaceticus schinkii, Clostridium ultunense, and Tepidanaerobacter acetatoxydans that comprise the formyltetrahydrofolate synthetase gene (fhs), encoding a key enzyme of the Wood-Ljungdahl pathway. We also discovered that the latter two harbor two fhs alleles. The fhs genes are phylogenetically separated and in the case of S. schinkii functionally linked to sulfate reducers. The T. acetatoxydans fhs1 cluster combines features of acetogens, sulfate reducers, and carbon monoxide oxidizers and is organized as a putative operon. The T. acetatoxydans fhs2 cluster encodes Wood-Ljungdahl pathway enzymes, which are also known to be involved in C1 carbon metabolism. Isolation of the enzymes illustrated that both formyltetrahydrofolate synthetases of T. acetatoxydans were functionally active. However, only fhs1 was expressed, confirming bidirectional usage of the pathway.