Physarum nitric oxide synthases: genomic structures and enzymology of recombinant proteins.
Physarum polycephalum expresses two closely related, calcium-independent NOSs (nitric oxide synthases). In our previous work, we showed that both NOSs are induced during starvation and apparently play a functional role in sporulation. In the present study, we characterized the genomic structures of both Physarum NOSs, expressed both enzymes recombinantly in bacteria and characterized their biochemical properties. Whereas the overall genomic organization of Physarum NOS genes is comparable with various animal NOSs, none of the exon-intron boundaries are conserved. Recombinant expression of clones with various N-termini identified N-terminal amino acids essential for enzyme activity, but not required for haem binding or dimerization, and suggests the usage of non-AUG start codons for Physarum NOSs. Biochemical characterization of the two Physarum isoenzymes revealed different affinities for L-arginine, FMN and 6R-5,6,7,8-tetrahydro-L-biopterin.
Messner
Simon
S
Division of Biological Chemistry, Biocenter, Innsbruck Medical University, Fritz-Pregl-Strasse 3/VI, Innsbruck, Austria.
Leitner
Stephan
S
Bommassar
Christian
C
Golderer
Georg
G
Gröbner
Peter
P
Werner
Ernst R
ER
Werner-Felmayer
Gabriele
G
D000818:Animals; D001120:Arginine / Q000378:metabolism; D001483:Base Sequence; D001708:Biopterins / Q000031:analogs & derivatives / Q000378:metabolism; D005486:Flavin Mononucleotide / Q000378:metabolism; D007527:Isoenzymes / Q000378:metabolism; D008969:Molecular Sequence Data; D019001:Nitric Oxide Synthase / Q000378:metabolism*; D016839:Physarum polycephalum / Q000201:enzymology* / Q000235:genetics; D012333:RNA, Messenger / Q000378:metabolism
D007527:Isoenzymes; D012333:RNA, Messenger; D001708:Biopterins; D005486:Flavin Mononucleotide; D001120:Arginine; D019001:Nitric Oxide Synthase; C003402:sapropterin