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Design and Characterization of Stabilized Derivatives of Human CD4D12 and CD4D1

Saha, Piyali and Barua, Bipasha and Bhattacharyya, Sanchari and Balamurali, MM and Schief, William R and Baker, David and Varadarajan, Raghavan (2011) Design and Characterization of Stabilized Derivatives of Human CD4D12 and CD4D1. In: Biochemistry, 50 (37). pp. 7891-7900.

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Official URL: http://pubs.acs.org/doi/abs/10.1021/bi200870r

Abstract

CD4 is present on the surface of T-lymphocytes and is the primary cellular receptor for HIV-1. CD4 consists of a cytoplasmic tail, one transmembrane region, and four extracellular domains, D1-D4. A construct consisting of the first two domains of CD4 (CD4D12) is folded and binds gp120 with similar affinity as soluble 4-domain CD4 (sCD4). However, the first domain alone (CD4D1) was previously shown to be largely unfolded and had 3-fold weaker affinity for gp120 when compared to sCD4 [Sharma, D.; et al. (2005) Biochemistry 44, 16192-16202]. We now report the design and characterization of three single-site mutants of CD4D12 (G6A, L51I, and V86L) and one multisite mutant of CD4D1 (G6A/L511/L5K/F98T). G6A, L51I, and V86L are cavity-filling mutations while L5K and F98T are surface mutations which were introduced to minimize the aggregation of CD4D1 upon removal of the second domain. Two mutations, G6A and V86L in CD4D12 increased the stability and yield of the protein relative to the wild-type protein. The mutant CD4D1 (CD4D1a) with the 4 mutations was folded and more stable compared to the original CD4D1, but both bound gp120 with comparable affinity. In in vitro neutralization assays, both CD4D1a and G6A-CD4D12 were able to neutralize diverse HIV-1 viruses with similar IC(50)s as 4-domain CD4. These stabilized derivatives of human CD4 can be useful starting points for the design of other more complex viral entry inhibitors.

Item Type: Journal Article
Additional Information: Copyright of this article belongs to American Chemical Society.
Department/Centre: Division of Biological Sciences > Molecular Biophysics Unit
Date Deposited: 28 Sep 2011 10:50
Last Modified: 28 Sep 2011 10:50
URI: http://eprints.iisc.ernet.in/id/eprint/40762

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