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Pepstatin Pull-Down at High pH Is a Powerful Tool for Detection and Analysis of Napsin A

Pepstatin Pull-Down at High pH Is a Powerful Tool for Detection and Analysis of Napsin A

Andreas Maurer, Hubert Kalbacher

Biochem Biophys Res Commun. 2019 Jul 12;515(1):145-148.

PMID: 31130231

Abstract:

Napsin A is an intracellular aspartic protease and biomarker of various malignancies like lung adenocarcinoma and ovarian clear cell carcinoma, but its detection is usually limited to immunohistochemical techniques gaining excellent information on its distribution but missing information about posttranslational modifications (e.g. maturation state) of the protein. We present a protocol for specific enrichment of napsin A from clinical or biological specimens, that facilitates detailed analysis of the protein. By using the exceptionally broad pH range under which napsin A binds to its inhibitor pepstatin A we achieve highly selective binding of napsin A while other aspartic proteases have negligible affinity. Using this method we demonstrate that lung napsin A in many mammals is a heterogeneous enzyme with a characteristic ladder-like appearance in SDS-PAGE that might be caused by proteolytically processed N- and/or C-termini, in contrast to the more homogeneous form found in kidneys and primary lung adenocarcinoma.

Chemicals Related in the Paper:

Catalog Number	Product Name	Structure	CAS Number	Price
AP26305033	Pepstatin A		26305-03-3	Price

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