Expression, Purification, and Activity of Putative Intramembrane Aspartyl Proteases From Diverse Species
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Thomas, Gwendell Michelle
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Abstract
Intramembrane aspartyl proteases (IAPs) cleave peptide bonds within the hydrophobic lipid membrane. The best characterized IAP is presenilin, the catalytic subunit of γ-Secretase, which is known for cleaving amyloid precursor protein into the amyloid-β peptide that aggregates in the brains of Alzheimer’s patients. More than 100 substrates have been documented for γ-Secretase, yet no consensus recognition sequence has emerged, at least in part due to the technical complexities of studying this membrane-embedded proteolytic system. To date, the presenilin homolog from Methanoculleus marisnigri JR1 (MCMJR1) has been the sole microbial model of non-eukaryotic IAPs for in vitro molecular studies. A recent bioinformatic study uncovered over 1000 putative IAP sequences lurking in archaeal and bacterial organisms. Here, we report recombinant expression, purification, and enzymatic activity of selected new IAPs. These new IAPs express well and are from noted archaea such as H. volcanii and Lokiarchaeota. These putative IAP orthologs share key structural and cleavage preference similarities with MCMJR1 IAP and presenilin. By studying the molecular biochemistry of more IAP family members, additional trends and insights regarding cleavage preferences and peculiarities will emerge. Such knowledge will further illuminate the fascinating fundamental and complex chemistry occurring within the lipid membrane.
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2022-04-27
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