Structural Basis for Nucleotide Hydrolysis by the Acid Sphingomyelinase-like Phosphodiesterase SMPDL3A

J Biol Chem. 2016 Mar 18;291(12):6376-85. doi: 10.1074/jbc.M115.711085. Epub 2016 Jan 20.


Sphingomyelin phosphodiesterase, acid-like 3A (SMPDL3A) is a member of a small family of proteins founded by the well characterized lysosomal enzyme, acid sphingomyelinase (ASMase). ASMase converts sphingomyelin into the signaling lipid, ceramide. It was recently discovered that, in contrast to ASMase, SMPDL3A is inactive against sphingomyelin and, surprisingly, can instead hydrolyze nucleoside diphosphates and triphosphates, which may play a role in purinergic signaling. As none of the ASMase-like proteins has been structurally characterized to date, the molecular basis for their substrate preferences is unknown. Here we report crystal structures of murine SMPDL3A, which represent the first structures of an ASMase-like protein. The catalytic domain consists of a central mixed β-sandwich surrounded by α-helices. Additionally, SMPDL3A possesses a unique C-terminal domain formed from a cluster of four α-helices that appears to distinguish this protein family from other phosphoesterases. We show that SMDPL3A is a di-zinc-dependent enzyme with an active site configuration that suggests a mechanism of phosphodiester hydrolysis by a metal-activated water molecule and protonation of the leaving group by a histidine residue. Co-crystal structures of SMPDL3A with AMP and α,β-methylene ADP (AMPCP) reveal that the substrate binding site accommodates nucleotides by establishing interactions with their base, sugar, and phosphate moieties, with the latter the major contributor to binding affinity. Our study provides the structural basis for SMPDL3A substrate specificity and sheds new light on the function of ASMase-like proteins.

Keywords: SMPDL3A; crystal structure; enzyme catalysis; nucleoside/nucleotide metabolism; sphingomyelinase; zinc.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, Non-P.H.S.

MeSH terms

  • Adenosine Monophosphate / chemistry*
  • Adenosine Triphosphate / chemistry
  • Amino Acid Sequence
  • Animals
  • Biocatalysis
  • Catalytic Domain
  • Cell Line
  • Crystallography, X-Ray
  • Humans
  • Hydrolysis
  • Kinetics
  • Mice
  • Models, Molecular
  • Molecular Sequence Data
  • Protein Binding
  • Protein Structure, Secondary
  • Sphingomyelin Phosphodiesterase / chemistry*
  • Substrate Specificity
  • Zinc / chemistry


  • Adenosine Monophosphate
  • Adenosine Triphosphate
  • SMPDL3A protein, mouse
  • Sphingomyelin Phosphodiesterase
  • Zinc

Associated data

  • PDB/1WAR
  • PDB/3LL8
  • PDB/5FC1
  • PDB/5FC5
  • PDB/5FC6
  • PDB/5FC7
  • PDB/5FCA
  • PDB/5FCB