T. Happe et al., Transcriptional and mutational analysis of the uptake hydrogenase of the filamentous cyanobacterium Anabaena variabilis ATCC 29413, J BACT, 182(6), 2000, pp. 1624-1631
A 10-kb DNA region of the cyanobacterium Anabaena variabilis ATCC 29413 con
taining the structural genes of the uptake hydrogenase (hupSL) was cloned a
nd sequenced. In contrast to the hupL, gene of Anabaena sp. strain PCC 7120
, which is interrupted by a 10.5-kb DNA fragment in vegetative cells, there
is no programmed rearrangement within the hupL gene during the heterocyst
differentiation of A. variabilis, The hupSL genes were transcribed as a 2.7
-kb operon and were induced only under nitrogen-fixing conditions, as shown
by Northern blot experiments and reverse transcriptase PCR. Primer extensi
on experiments with a fluorescence-labeled oligonucleotide primer confirmed
these results and identified the 5' start of the mRNA transcript 103 bp up
stream of the ATG initiation codon, A consensus sequence in the promoter th
at is recognized by the fumarate nitrate reductase regulator (Fnr) could be
detected. The hupSL operon in A. variabilis was interrupted by an interpos
on deletion (mutant strain AVM13). Under N-2-fixing conditions, the mutant
strain exhibited significantly increased rates in H-2 accumulation and prod
uced three times more hydrogen than the wild type. These results indicate t
hat the uptake hydrogenase is catalytically active in the wild type and tha
t the enzyme reoxidizes the H-2 developed by the nitrogenase, The Nh phenot
ype of the mutant strain showed a slight decrease of acetylene reduction co
mpared to that of the wild type.