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J Biol Chem. 2015 Apr 24;290(17):10972-80. doi: 10.1074/jbc.M115.642413. Epub 2015 Feb 26.

From Protease to Decarboxylase: THE MOLECULAR METAMORPHOSIS OF PHOSPHATIDYLSERINE DECARBOXYLASE.

Author information

1
From the Department of Medicine, Basic Science Section, National Jewish Health, Denver, Colorado 80206.
2
the Department of Immunology and Infectious Diseases, Harvard School of Public Health, Boston, Massachusetts 02115.
3
the Department of Internal Medicine, Section of Infectious Diseases, Yale School of Medicine, New Haven, Connecticut 06520, and.
4
From the Department of Medicine, Basic Science Section, National Jewish Health, Denver, Colorado 80206, voelkerd@njhealth.org.

Abstract

Phosphatidylserine decarboxylase (PSDs) play a central role in the synthesis of phosphatidylethanolamine in numerous species of prokaryotes and eukaryotes. PSDs are unusual decarboxylase containing a pyruvoyl prosthetic group within the active site. The covalently attached pyruvoyl moiety is formed in a concerted reaction when the PSD proenzyme undergoes an endoproteolytic cleavage into a large β-subunit, and a smaller α-subunit, which harbors the prosthetic group at its N terminus. The mechanism of PSD proenzyme cleavage has long been unclear. Using a coupled in vitro transcription/translation system with the soluble Plasmodium knowlesi enzyme (PkPSD), we demonstrate that the post-translational processing is inhibited by the serine protease inhibitor, phenylmethylsulfonyl fluoride. Comparison of PSD sequences across multiple phyla reveals a uniquely conserved aspartic acid within an FFXRX6RX12PXD motif, two uniquely conserved histidine residues within a PXXYHXXHXP motif, and a uniquely conserved serine residue within a GS(S/T) motif, suggesting that PSDs belong to the D-H-S serine protease family. The function of the conserved D-H-S residues was probed using site-directed mutagenesis of PkPSD. The results from these mutagenesis experiments reveal that Asp-139, His-198, and Ser-308 are all essential for endoproteolytic processing of PkPSD, which occurs in cis. In addition, within the GS(S/T) motif found in all PSDs, the Gly-307 residue is also essential, but the Ser/Thr-309 is non-essential. These results define the mechanism whereby PSDs begin their biochemical existence as proteases that execute one autoendoproteolytic cleavage reaction to give rise to a mature PSD harboring a pyruvoyl prosthetic group.

KEYWORDS:

Autoendoproteolysis; Membrane; Phosphatidylserine Decarboxylase; Phospholipid; Plasmodium; Protease; Pyruvoyl-enzyme

PMID:
25724650
PMCID:
PMC4409258
DOI:
10.1074/jbc.M115.642413
[Indexed for MEDLINE]
Free PMC Article

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