Abstract
BackgroundMycoplasma suis belongs to a group of highly specialized hemotrophic bacteria that attach to the surface of host erythrocytes. Hemotrophic mycoplasmas are uncultivable and the genomes are not sequenced so far. Therefore, there is a need for the clarification of essential metabolic pathways which could be crucial barriers for the establishment of an in vitro cultivation system for these veterinary significant bacteria.Inorganic pyrophosphatases (PPase) are important enzymes that catalyze the hydrolysis of inorganic pyrophosphate PPi to inorganic phosphate Pi. PPases are essential and ubiquitous metal-dependent enzymes providing a thermodynamic pull for many biosynthetic reactions. Here, we describe the identification, recombinant production and characterization of the soluble (s)PPase of Mycoplasma suis.ResultsScreening of genomic M. suis libraries was used to identify a gene encoding the M. suis inorganic pyrophosphatase (sPPase). The M. suis sPPase consists of 164 amino acids with a molecular mass of 20 kDa. The highest identity of 63.7% was found to the M. penetrans sPPase. The typical 13 active site residues as well as the cation binding signature could be also identified in the M. suis sPPase. The activity of the M. suis enzyme was strongly dependent on Mg2+ and significantly lower in the presence of Mn2+ and Zn2+. Addition of Ca2+ and EDTA inhibited the M. suis sPPase activity. These characteristics confirmed the affiliation of the M. suis PPase to family I soluble PPases. The highest activity was determined at pH 9.0. In M. suis the sPPase builds tetramers of 80 kDa which were detected by convalescent sera from experimentally M. suis infected pigs.ConclusionThe identification and characterization of the sPPase of M. suis is an additional step towards the clarification of the metabolism of hemotrophic mycoplasmas and, thus, important for the establishment of an in vitro cultivation system. As an antigenic and conserved protein the M. suis sPPase could in future be further analyzed as a diagnostic antigen.
Highlights
Mycoplasma suis belongs to a group of highly specialized hemotrophic bacteria that attach to the surface of host erythrocytes
Mycoplasma suis belongs to a group of highly specialized uncultivable hemotrophic bacteria within the family Mycoplasmataceae that attach to the surface of host erythrocytes [1,2]
Identification of the molecular weight standard; (Ms). suis inorganic pyrophosphatase (PPase) The soluble inorganic pyrophoshatase (sPPase) of M. suis was identified by screening of genomic libraries of M. suis using shot gun sequencing
Summary
Mycoplasma suis belongs to a group of highly specialized hemotrophic bacteria that attach to the surface of host erythrocytes. Mycoplasma suis belongs to a group of highly specialized uncultivable hemotrophic bacteria within the family Mycoplasmataceae that attach to the surface of host erythrocytes [1,2]. Inorganic pyrophosphate (PPi) is an important by-product of many biosynthetic processes, and sPPases which hydrolyze PPi to inorganic phosphate (Pi), are essential and ubiquitous metal-dependent enzymes providing a thermodynamic pull for many biosynthetic reactions [11,12,13]. Characterization of essential enzymes in the metabolism of hemotrophic mycoplasmas are important steps towards the establishment of an in vitro cultivation system for this group of hitherto uncultivable hemotrophic bacteria
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