Genes encoding the candidate enzyme for anaerobic activation of n-alkanes in the denitrifying bacterium, strain HxN1.
Strain HxN1, a member of the Betaproteobacteria, can grow anaerobically by denitrification with n-alkanes. n-Alkanes are apparently activated by subterminal carbon addition to fumarate yielding (1-methylalkyl)succinates, the postulated enzyme being (1-methylalkyl)succinate synthase (Mas). Genes encoding this enzyme (mas) were searched for via proteins that were specifically formed in n-hexane-grown cells (in comparison with caproate-grown cells), as revealed by two-dimensional gel electrophoresis. Partial amino acid sequencing and subsequent probe development for hybridization of restricted DNA led to the identification of a gene cluster. Deduced proteins are similar to the subunits of benzylsuccinate synthase (Bss), the toluene-activating enzyme in other anaerobic bacteria and its activase. The tentative (1-methylalkyl)succinate synthase is presumably a heterotrimer (MasDEC) which, like benzylsuccinate synthase, contains a motif (in MasD, the large subunit) characteristic of glycyl radical-bearing sites. Based on amino acid sequence comparison, the tentative (1-methylalkyl)succinate synthase branches outside of the phylogenetic cluster of benzylsuccinate synthases from different organisms and represents a separate line of descent within glycyl radical enzymes. n-Hexane-induced co-transcription of the mas genes and additional genes of an apparent operon was demonstrated by Northern hybridization experiments.
Grundmann
Olav
O
Max-Planck-Institut für Marine Mikrobiologie, Celsiusstrasse 1, D-28359 Bremen, Germany. ogrundma@mpi-bremen.de
Behrends
Astrid
A
Rabus
Ralf
R
Amann
Judith
J
Halder
Thomas
T
Heider
Johann
J
Widdel
Friedrich
F
D000473:Alkanes / Q000378:metabolism*; D000693:Anaerobiosis; D001426:Bacterial Proteins / Q000737:chemistry / Q000235:genetics / Q000378:metabolism; D001483:Base Sequence; D020562:Betaproteobacteria / Q000201:enzymology* / Q000235:genetics / Q000254:growth & development; D019755:Carbon-Carbon Lyases / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D015180:Electrophoresis, Gel, Two-Dimensional; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D006586:Hexanes / Q000378:metabolism; D008969:Molecular Sequence Data; D005810:Multigene Family; D009573:Nitrites / Q000378:metabolism*; D017422:Sequence Analysis, DNA; D013386:Succinates / Q000378:metabolism*
D000473:Alkanes; D001426:Bacterial Proteins; D006586:Hexanes; D009573:Nitrites; D013386:Succinates; D019755:Carbon-Carbon Lyases