**An Engineered Microvirin Variant with Identical Structural Domains Potently Inhibits Human Immunodeficiency Virus and Hepatitis C Virus Cellular Entry**

**Munazza Shahid 1,**†**, Amina Qadir 1,**†**, Jaewon Yang 2, Izaz Ahmad 1, Hina Zahid 1, Shaper Mirza 1, Marc P. Windisch 2,3 and Syed Shahzad-ul-Hussan 1,\***


Received: 13 January 2020; Accepted: 22 January 2020; Published: 11 February 2020

**Abstract:** Microvirin (MVN) is one of the human immunodeficiency virus (HIV-1) entry inhibitor lectins, which consists of two structural domains sharing 35% sequence identity and contrary to many other antiviral lectins, it exists as a monomer. In this study, we engineered an MVN variant, LUMS1, consisting of two domains with 100% sequence identity, thereby reducing the chemical heterogeneity, which is a major factor in eliciting immunogenicity. We determined carbohydrate binding of LUMS1 through NMR chemical shift perturbation and tested its anti-HIV activity in single-round infectivity assay and its anti-hepatitis C virus (HCV) activity in three different assays including HCVcc, HCVpp, and replicon assays. We further investigated the effect of LUMS1 on the activation of T helper (Th) and B cells through flow cytometry. LUMS1 showed binding to α(1-2)mannobiose, the minimum glycan epitope of MVN, potently inhibited HIV-1 and HCV with EC50 of 37.2 and 45.3 nM, respectively, and showed negligible cytotoxicity with CC50 > 10 μM against PBMCs, Huh-7.5 and HepG2 cells, and 4.9 μM against TZM-bl cells. LUMS1 did not activate Th cells, and its stimulatory effect on B cells was markedly less as compared to MVN. Together, with these effects, LUMS1 represents a potential candidate for the development of antiviral therapies.

**Keywords:** microvirin; lectin; human immunodeficiency virus; hepatitis C virus; antiviral inhibitor; non-immunogenic; viral entry; protein drugs; LUMS1
