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Issue 19, 2021
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Development of a RPLC-UV method for monitoring uncleaved HIV-1 envelope glycoprotein

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Abstract

One of the HIV-1 vaccine design efforts has focused on developing a recombinant HIV-1 trimeric envelope glycoprotein (Env) as an immunogen to induce broadly neutralizing antibodies. A native-like immunogen, the BG505.DS.SOSIP.664 gp140 (Env) construct has been well-characterized as a vaccine candidate. This vaccine candidate comprises of three identical gp120 and truncated gp41 subunits that form into a trimer of heterodimers. During production, recombinant Env is expressed as a gp140 precursor polypeptide in which a furin cleavable site is engineered to generate a heterodimer of gp120 and gp41 subunits. Each heterodimer is connected by an intermolecular disulfide bond, and three heterodimers form into a trimer. Furin cleavage is an important factor to mimic native-like HIV-1 Env conformations and is needed to help induce an immune response. Therefore, it is critical to monitor cleavage for ensuring functionality of the Env vaccine product. In this paper, a new RPLC-UV method coupled with reduction was developed to routinely determine the percentage of uncleaved gp140 relative to the cleaved gp120 and gp41 subunits. Baseline separation was achieved among the gp120, gp41 and uncleaved gp140 peaks, thus enabling relative quantification of uncleaved gp140. Overall, this RPLC-UV approach has been successfully applied to support Env vaccine candidate developments.

Graphical abstract: Development of a RPLC-UV method for monitoring uncleaved HIV-1 envelope glycoprotein

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Supplementary files

Article information


Submitted
11 Jan 2021
Accepted
27 Apr 2021
First published
06 May 2021

Anal. Methods, 2021,13, 2183-2188
Article type
Communication

Development of a RPLC-UV method for monitoring uncleaved HIV-1 envelope glycoprotein

N. A. Schneck, A. L. Vinitsky, V. B. Ivleva, X. Wang, D. B. Gowetski and Q. P. Lei, Anal. Methods, 2021, 13, 2183
DOI: 10.1039/D1AY00072A

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