# Comprehensive Insights into the Molecular Basis of HIV Glycoprotein
## 2 Proteins Engaged in HIV Entry
### Env/gp160
#### 2.1.1 Synthesis and Trafficking of Env/gp160
Initially, Env is synthesized and glycosylated within the rough endoplasmic reticulum (RER) as a 160-kDa precursor protein, designated gp160. This precursor protein predominantly oligomerizes into trimers. Concurrently, the precursor Gag protein (Pr55 Gag) is synthesized on cytosolic ribosomes and directed to the plasma membrane (PM), where it multimerizes within lipid rafts (not depicted here) to form nascent virus particles. The oligomerized gp160 is subsequently transported to the Golgi apparatus and the trans-Golgi network (TGN). Here, gp160 undergoes proteolytic processing to generate the mature surface glycoprotein gp120 and the transmembrane glycoprotein gp41. The gp120/gp41 complexes proceed through the secretory pathway, eventually reaching the plasma membrane, where they are incorporated into virus particles as trimeric spikes. At the plasma membrane, Env can undergo endocytosis mediated by clathrin adaptor complexes, leading to its internalization into early endosomes (EE). Within the cell, internalized Env has two potential fates: it can be routed to late endosomes/multivesicular bodies (LE/MVBs) for subsequent degradation in lysosomes, or it can be recycled back to the plasma membrane via recycling endosomes. The domains of Gag and Env are detailed in the inset located at the top left of the diagram. Reprinted (adapted) with permission from [31; 56] Copyright Elsevier (2024). Figure 2: HIV-1 gp160 protein structure and post-translational modifications. (**a**) Structure and processing of precursor gp160. The precursor gp160 protein of HIV-1 includes a signal peptide (SP) that is cleaved during translation. Post-translationally, gp160 is processed into the surface subunit (gp120) and the transmembrane subunit (gp41) within the Golgi complex at a specific furin cleavage site. The gp120 subunit comprises five variable domains (V1–V5) and five constant domains (C1–C5). In contrast, the gp41 subunit features extracellular constant domains (C6 and C7) that includes the fusion peptide (FP), hept repeats (HR1 and HR2), the membrane-proximal external region (MPER), a transmembrane domain (TMD), and a cytoplasmic tail (CT). An enlarged depiction of the gp41 CT highlights several critical motifs: the internalization signal YSPL, the Kennedy sequence, the amphipathic \(\alpha\)-helices LLP-1, LLP-2 and LLP-3, and a C-terminal dileucine motif (LL) implicated in the endocytosis and intracellular distribution of Env. The glycosylation sites on various HIV-1 gp120 variants are shown in a structure-based sequence alignment. These sites are called complex or high-mannose glycosylation sites. The domains are color-coded, with the N and C termini indicated. Disulfide bonds are represented as red lines, and the inner-domain \(\beta\) sandwich is boxed. Oligomannose and complex glycans are symbolized by three or two-pronged forked symbols, respectively. Reprinted (adapted) from [34; 57; 58] under a Creative Commons license. (**b**) Conformational changes during gp120 folding and signal-peptide cleavage. Stage I: Upon the completion of translation, gp120 remains largely unfolded. An \(\alpha\)-helical structure around the signal peptide impedes the cleavage of the signal peptide. Stage II: As long as the signal peptide is attached, the cysteine at position 28 (C28) promotes intramolecular disulfide isomerization by interacting with downstream cysteine residues. The tethering of the N-terminus restricts conformational freedom, potentially aiding the folding of the N-terminal region. Stage III: The folding and integration of the inner-domain \(\beta\) sandwich disrupt the helical structure of the signal peptide, exposing the consensus cleavage site. Stage IV: The cleavage of the signal peptide stabilizes the conformation of gp120 by removing the free sulfhydryl group of C28, thus halting further disulfide isomerization. In the schematic, the inner domain is shaded in gray, the outer domain is pink, the variable loops are green, and the signal peptide is orange.