Protective Mechanisms against Homocysteine Toxicity

Abstract

Homocysteine (Hcy) editing by methionyl-tRNA synthetase results in the formation of Hcy-thiolactone and initiates a pathway that has been implicated in human disease. In addition to being cleared from the circulation by urinary excretion, Hcythiolactone is detoxified by the serum Hcy-thiolactonase/paraoxonase carried on high density lipoprotein. Whether Hcy-thiolactone is detoxified inside cells was unknown. Here we show that Hcy-thiolactone is hydrolyzed by an intracellular enzyme, which we have purified to homogeneity from human placenta and identified by proteomic analyses as human bleomycin hydrolase (hBLH). We have also purified an Hcy-thiolactonase from the yeast Saccharomyces cerevisiae and identified it as yeast bleomycin hydrolase (yBLH). BLH belongs to a family of evolutionarily conserved cysteine aminopeptidases, and its only known biologically relevant function was deamidation of the anticancer drug bleomycin. Recombinant hBLH or yBLH, expressed in Escherichia coli, exhibits Hcy-thiolactonase activity similar to that of the native enzymes. Active site mutations, C73A for hBLH and H369A for yBLH, inactivate Hcy-thiolactonase activities. Yeast blh1 mutants are deficient in Hcy-thiolactonase activity in vitro and in vivo, produce more Hcy-thiolactone, and exhibit greater sensitivity to Hcy toxicity than wild type yeast cells. Our data suggest that BLH protects cells against Hcy toxicity by hydrolyzing intracellular Hcy-thiolactone.

Neutral Cysteine Protease Bleomycin Hydrolase Is Essential for the Breakdown of Deiminated Filaggrin into Amino Acids

“…However, this function has remained enigmatic for many years. Recently, BH was shown to be involved in the post-translational processing of peptides for antigen presentation (36) and a protective mechanism against homocysteine toxicity (37). Furthermore, BH has been shown to interact with several proteins, such as cAMP-binding ectoprotein in yeast (38), the human homologue of ubiquitin-conjugating enzyme 9 (39), human ribosomal proteins (40), and amyloid precursor protein (41).…”

Bleomycin Hydrolase Is Regulated Biphasically in a Differentiation- and Cytokine-dependent Manner

“…BH exhibits mainly broad specificity aminopeptidase activity, with citrulline being the most effective substrate (12,14). It is involved in homocysteinethiolactone metabolism (17), antigen presentation (18,19), and amyloid-␤-peptide clearance (20,21). Interestingly, BH knockout mouse shows analogous phenotypes to those of filaggrindeficient flaky tail ft/ft mouse, including ichthyosis and atopylike phenotype and tail constriction (22)(23)(24).…”

The Lap3p aminopeptidase is not solely responsible for bleomycin resistance in Candida albicans

“…Numerous cellular functions other than bleomycin detoxification have been described for yeast Lap3p and its orthologs (Koldamova et al, 1998;Montoya et al, 1998;Zimny et al, 2006;Montoya et al, 2007;Kamata et al, 2011) Furthermore, cysteine proteases throughout nature have demonstrated a role in a large variety of processes, ranging from signaling pathways in many organisms to programmed cell death (Degterev et al, 2003). Our results presented here raise further questions regarding the nature of the function of the Candida protein.…”

“…Although GAL6 deletion mutants are viable, Gal6p (Lap3p) is known to exert a modest negative effect on GAL promoter mediated transcription (Zheng et al, 1997). In humans, BLH1 is expressed in most tissue types (Takeda et al, 1996), and functions in protection against homocysteine toxicity (Zimny et al, 2006) and progression of atopic dermatitis. A correlative association with development of astrogliosis has also been attributed to abnormalities in BLH1 function (Montoya et al, 1998;Montoya et al, 2007).…”