Mass spectrometric evidence for neuropeptide-amidating enzymes in Caenorhabditis elegans

• Published in: The Journal of biological chemistry Vol. 293; no. 16; pp. 6052 - 6063
• Main Authors: Van Bael, Sven, Watteyne, Jan, Boonen, Kurt, De Haes, Wouter, Menschaert, Gerben, Ringstad, Niels, Horvitz, H. Robert, Schoofs, Liliane, Husson, Steven J., Temmerman, Liesbet
• Format: Journal Article
• Language: English
• Published: United States Elsevier Inc 20.04.2018; American Society for Biochemistry and Molecular Biology
• Subjects: amidation; Amidine-Lyases - chemistry; Amidine-Lyases - genetics; Amidine-Lyases - metabolism; Amino Acid Sequence; Animals; Biosynthetic Pathways; Caenorhabditis elegans (C. elegans); Caenorhabditis elegans - chemistry; Caenorhabditis elegans - genetics; Caenorhabditis elegans - metabolism; Caenorhabditis elegans Proteins - chemistry; Caenorhabditis elegans Proteins - genetics; Caenorhabditis elegans Proteins - metabolism; Copper - metabolism; copper monooxygenase; Gene Deletion; Genomics and Proteomics; Humans; mass spectrometry (MS); Mixed Function Oxygenases - chemistry; Mixed Function Oxygenases - genetics; Mixed Function Oxygenases - metabolism; Multienzyme Complexes - chemistry; Multienzyme Complexes - genetics; Multienzyme Complexes - metabolism; Mutation; neuropeptide; Neuropeptides - genetics; Neuropeptides - metabolism; PAL; PAM; peptides; peptidomics; PHM; Sequence Alignment; Tandem Mass Spectrometry; amidation; PHM; peptidomics; neuropeptide; Caenorhabditis elegans (C. elegans); peptides; copper monooxygenase; PAL; PAM; mass spectrometry (MS)
• ISSN: 0021-9258; 1083-351X
• DOI: 10.1074/jbc.RA117.000731

Neuropeptides constitute a vast and functionally diverse family of neurochemical signaling molecules and are widely involved in the regulation of various physiological processes. The nematode Caenorhabditis elegans is well-suited for the study of neuropeptide biochemistry and function, as neuropeptide biosynthesis enzymes are not essential for C. elegans viability. This permits the study of neuropeptide biosynthesis in mutants lacking certain neuropeptide-processing enzymes. Mass spectrometry has been used to study the effects of proprotein convertase and carboxypeptidase mutations on proteolytic processing of neuropeptide precursors and on the peptidome in C. elegans. However, the enzymes required for the last step in the production of many bioactive peptides, the carboxyl-terminal amidation reaction, have not been characterized in this manner. Here, we describe three genes that encode homologs of neuropeptide amidation enzymes in C. elegans and used tandem LC-MS to compare neuropeptides in WT animals with those in newly generated mutants for these putative amidation enzymes. We report that mutants lacking both a functional peptidylglycine α-hydroxylating monooxygenase and a peptidylglycine α-amidating monooxygenase had a severely altered neuropeptide profile and also a decreased number of offspring. Interestingly, single mutants of the amidation enzymes still expressed some fully processed amidated neuropeptides, indicating the existence of a redundant amidation mechanism in C. elegans. All MS data are available via ProteomeXchange with the identifier PXD008942. In summary, the key steps in neuropeptide processing in C. elegans seem to be executed by redundant enzymes, and loss of these enzymes severely affects brood size, supporting the need of amidated peptides for C. elegans reproduction.