Characterization of Rhizobium japonicum hydrogen uptake genes

Recombinant cosmids from a gene library of the DNA from Hup+ Rhizobium japonicum 122DES previously have been shown to restore hydrogenase activity when transferred by conjugation into certain Hup- mutants of R. japonicum. We generated a restriction map covering 32.2 kilobases of this cosmid DNA. At least 25.3 kilobases of the cosmid pHU1 were shown to have the same arrangement as those in the genome of strain 122DES. Analysis of Tn5 insertions into the 122DES genome indicates that hup-specific sequences occur in a region spanning about 15 kilobases of insert DNA within pHU1. Introduction of pHU1 into five out of six R. japonicum Hup- mutants resulted in a Hup+ phenotype in some transconjugants. Three of the mutations appear to be in transcriptional units completely contained within pHU1, whereas the other two must be in genes that are at least partially contained within pHU1. pBR235 derivatives containing fragments of hup DNA can be transferred into the R. japonicum Hup- mutant PJ18nal if the derivatives contain a region of homology with the R. japonicum genome. The hup mutation in strain PJ18nal appears to be dominant. The hup genes in R. japonicum strain 122DES appear to be organized in at least two, and probably three, transcriptional units.