N-acyl Taurines Research Articles

A FAAH-Regulated Class of N-Acyl Taurines That Activates TRP Ion Channels

Fatty acid amide hydrolase (FAAH) is an integral membrane enzyme that catabolizes several bioactive lipids in vivo. Most of the physiological substrates of FAAH characterized to date belong to the N-acyl ethanolamine (NAE) class of fatty acid amides, including the endocannabinoid anandamide, the anti-inflammatory lipid N-palmitoyl ethanolamine, and the satiating factor N-oleoyl ethanolamine. We recently identified a second structural class of fatty acid amides regulated by FAAH in vivo: the N-acyl taurines (NATs). Global metabolite profiling revealed high concentrations of long chain (> or = C20) saturated NATs in the central nervous system (CNS) of FAAH(-/-) mice. Here, we use metabolite profiling to characterize the FAAH-NAT system in peripheral mouse tissues. Livers and kidneys of FAAH(-/-) mice possessed dramatic elevations in NATs, which, in contrast to those detected in the CNS, were enriched in polyunsaturated acyl chains (e.g., C20:4, C22:6). Peripheral NATs rose more than 10-fold within 1 h following pharmacological inactivation of FAAH and reached levels up to approximately 5000 pmol/g tissue (C22:6 in kidney), implicating a constitutive and highly active pathway for NAT metabolism in which FAAH plays an integral part. Interestingly, NATs were found to activate multiple members of the transient receptor potential (TRP) family of calcium channels, including TRPV1 and TRPV4, which are both expressed in kidney. The dramatic elevation in endogenous levels of NATs following acute or chronic inactivation of FAAH, in conjunction with the pharmacological effects of these lipids on TRP channels, suggests the existence of a second major lipid signaling system regulated by FAAH in vivo.

Structure-Based Design of a FAAH Variant That Discriminates between the N-Acyl Ethanolamine and Taurine Families of Signaling Lipids

Fatty acid amide hydrolase (FAAH) inactivates a large and diverse class of endogenous signaling lipids termed fatty acid amides. Representative fatty acid amides include the N-acyl ethanolamines (NAEs) anandamide, which serves as an endogenous ligand for cannabinoid receptors, and N-oleoyl and N-palmitoyl ethanolamine, which produce satiety and anti-inflammatory effects, respectively. Global metabolite profiling studies of FAAH (-/-) mice have recently identified a second class of endogenous FAAH substrates: the N-acyl taurines (NATs). To determine the metabolic and signaling functions performed by NAEs and NATs in vivo, a FAAH variant that discriminates between these two substrate classes would be of value. Here, we report the structure-guided design of a point mutant in the active site of FAAH that selectively disrupts interactions with NATs. This glycine-to-aspartate (G268D) mutant was found to exhibit wild-type kinetic parameters with NAEs, but more than a 100-fold reduction in activity with NATs attributable to combined effects on Km and kcat values. These in vitro properties were also observed in living cells, where WT-FAAH and the G268D mutant displayed equivalent hydrolytic activity with NAEs, but the latter enzyme was severely impaired in its ability to catabolize NATs. The G268D FAAH mutant may thus serve as a valuable research tool to illuminate the unique roles played by the NAE and NAT classes of signaling lipids in vivo.

Discovery metabolite profiling — forging functional connections between the proteome and metabolome

Of primary interest for every enzyme is the identification of its physiological substrates. However, the vast structural diversity of endogenous metabolites, coupled with the overlapping activities of numerous enzymes, makes it difficult to deduce the identity of natural substrates for a given enzyme based on in vitro experiments. To address this challenge, we recently introduced an LC–MS based analytical method termed discovery metabolite profiling (DMP) to evaluate the global metabolic effects of enzyme inactivation in vivo. We have applied DMP to study mice lacking the enzyme fatty acid amide hydrolase (FAAH), which degrades the endocannabinoid family of signaling lipids. DMP identified several previously uncharacterized FAAH substrates, including a structurally novel class of brain lipids that represent conjugates of very long chain fatty acids with the amino acid derivative taurine [ N-acyl taurines (NATs)]. These findings show that DMP can establish direct connections between the proteome and metabolome and thus offers a powerful strategy to assign physiological functions to enzymes in the post-genomic era.