title: Identification and biochemical characterization of an acid sphingomyelinase-like protein from the bacterial plant pathogen Ralstonia solanacearum that hydrolyzes ATP to AMP but not sphingomyelin to ceramide.
abstract: Acid sphingomyelinase (aSMase) is a human enzyme that catalyzes the hydrolysis of sphingomyelin to generate the bioactive lipid ceramide and phosphocholine. ASMase deficiency is the underlying cause of the genetic diseases Niemann-Pick Type A and B and has been implicated in the onset and progression of a number of other human diseases including cancer, depression, liver, and cardiovascular disease. ASMase is the founding member of the aSMase protein superfamily, which is a subset of the metallophosphatase (MPP) superfamily. To date, MPPs that share sequence homology with aSMase, termed aSMase-like proteins, have been annotated and presumed to function as aSMases. However, none of these aSMase-like proteins have been biochemically characterized to verify this. Here we identify RsASML, previously annotated as RSp1609: acid sphingomyelinase-like phosphodiesterase, as the first bacterial aSMase-like protein from the deadly plant pathogen Ralstonia solanacearum based on sequence homology with the catalytic and C-terminal domains of human aSMase. A biochemical characterization of RsASML does not support a role in sphingomyelin hydrolysis but rather finds RsASML capable of acting as an ATP diphosphohydrolase, catalyzing the hydrolysis of ATP and ADP to AMP. In addition, RsASML displays a neutral, not acidic, pH optimum and prefers Ni2+ or Mn2+, not Zn2+, for catalysis. This alters the expectation that all aSMase-like proteins function as acid SMases and expands the substrate possibilities of this protein superfamily to include nucleotides. Overall, we conclude that sequence homology with human aSMase is not sufficient to predict substrate specificity, pH optimum for catalysis, or metal dependence. This may have implications to the biochemically uncharacterized human aSMase paralogs, aSMase-like 3a (aSML3a) and aSML3b, which have been implicated in cancer and kidney disease, respectively, and assumed to function as aSMases.
authors.affiliation: Department of Medicine and the Stony Brook University Cancer Center, Stony Brook University, Stony Brook, New York, United States of America.
authors.forename: Michael V
authors.initials: MV
authors.lastname: Airola
authors.affiliation: Department of Biochemistry and Molecular Biology, Medical University of South Carolina, Charleston, South Carolina, United States of America.
authors.forename: Jessica M
authors.initials: JM
authors.lastname: Tumolo
authors.affiliation: Department of Medicine and the Stony Brook University Cancer Center, Stony Brook University, Stony Brook, New York, United States of America.
authors.forename: Justin
authors.initials: J
authors.lastname: Snider
authors.affiliation: Department of Medicine and the Stony Brook University Cancer Center, Stony Brook University, Stony Brook, New York, United States of America.
authors.forename: Yusuf A
authors.initials: YA
authors.lastname: Hannun
mesh_terms: D000249:Adenosine Monophosphate / Q000737:chemistry* / Q000378:metabolism; D000255:Adenosine Triphosphate / Q000737:chemistry* / Q000378:metabolism; D001426:Bacterial Proteins / Q000737:chemistry* / Q000378:metabolism; D006801:Humans; D006868:Hydrolysis; D043368:Ralstonia solanacearum / Q000201:enzymology*; D013108:Sphingomyelin Phosphodiesterase / Q000737:chemistry* / Q000378:metabolism; D013109:Sphingomyelins / Q000737:chemistry / Q000378:metabolism; D013379:Substrate Specificity
chemical_list: D001426:Bacterial Proteins; D013109:Sphingomyelins; D000249:Adenosine Monophosphate; D000255:Adenosine Triphosphate; D013108:Sphingomyelin Phosphodiesterase