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Curr Chem Genomics. 2012;6:38-47. doi: 10.2174/1875397301206010038. Epub 2012 Sep 20.

Utilizing HaloTag Technology to Track the Fate of PCSK9 from Intracellular vs. Extracellular Sources.

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1
Department of Atherosclerosis, Merck Research Laboratories, Rahway, NJ 07065, USA.

Abstract

The function of a particular protein is dependent upon its localization and milieu. The ability to track the "fate" of a protein is a valuable tool to elucidate its function. We present the use of HaloTag technology to study the localization and fate of human Proprotein Convertase Subtilisin-like Kexin type 9 (PCSK9).The role of PCSK9 in the regulation of circulating low density lipoprotein-cholesterol (LDL-c) levels is ascribed to binding of circulating PCSK9 to the LDL receptor (LDLR) and subsequent lysosomal degradation of LDLR. However, hints in the literature indicate that intracellular PCSK9 may act on the LDLR, possibly during processing of newly synthesized protein. To address this question, the source and fate of intracellular PCSK9 requires further investigation.We applied HaloTag technology to distinguish the source of intracellular PCSK9 and showed that newly synthesized intracellular PCSK9 has unique localization from the PCSK9 after re-uptake. This suggests different functions of PCSK9 while interacting with the LDLR.

KEYWORDS:

Confocal imaging; HaloTag; LDL receptor; PCSK9; intracellular trafficking; protein label; protein uptake; subcellu-lar localization.

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