Biophysical properties of the isolated spike protein binding helix of human ACE2

Abstract

The entry of the SARS-CoV2 virus in human cells is mediated by the binding of its surface spike protein to the human Angiotensin-Converting Enzyme 2 (ACE2) receptor. A 23 residues long helical segment (SBP1) at the binding interface of human ACE2 interacts with viral spike protein and therefore, has generated considerable interest as a recognition element for virus detection. Unfortunately, emerging reports indicate that the affinity of SBP1 to the receptor-binding domain (RBD) of the spike protein is much lower than that of the ACE2 receptor itself. Here, we examine the biophysical properties of SBP1 to reveal factors leading to its low affinity for the spike protein. While SBP1 shows good solubility (solubility > 0.8 mM), CD spectroscopy shows that it is mostly disordered with some anti-parallel beta-sheet content, and no helicity. The helicity is substantial (> 20%) only upon adding high concentrations (≥ 20% v/v) of 2,2,2-trifluoroethanol, a helix-promoter. Fluorescence correlation spectroscopy and single molecule photobleaching studies show that the peptide oligomerizes at concentrations > 50 nM. We hypothesized that mutating the hydrophobic residues (F28, F32, and F40) of SBP1 which do not directly interact with the spike protein to alanine would reduce peptide oligomerization without affecting its spike binding affinity. While the mutant peptide (SBP1 mod ) shows substantially reduced oligomerization propensity, it does not show improved helicity. Our study shows that the failure of efforts so far to produce a short SBP1 mimic with a high affinity for the spike protein is not only due to the lack of helicity, but also due to the heretofore unrecognized problem of oligomerization.

Comparative study of SARS-CoV-2 infection in different cell types: Biophysical-computational approach to the role of potential receptors

“…Neuropilin 1 (NRP1), Basigin2 (CD147), FURIN protease, Transmembrane Serine 2 (TMPRSS2), Angiotensin Converting Enzyme 2 (ACE2), Protein of Thermal shock A5 (HSPA5), Angiotensin II receptor type 2 (AGTR2). Model 1, A549 (∼4.4x10 −2 Pa/s) [ 27 ]; Model 2, HeLa negative control (∼4.4x10 −2 Pa/s) [ 27 ]; Model 3, Normal Swiss 3T3 cells (∼2.4x10 −2 Pa/s) [ 27 , 28 ]; Model 4, ASTC-a-1 (∼1.6x10 −3 Pa/s) [ 29 ]; Model 5, H1299 (∼1.1x10 2 Pa/s) [ 30 ]; Model 6, standard viscosity of water (∼1.0x10 −3 Pa/s) [ 56 ]. *, time calculated in model 6.…”

Inhibition of Human Colorectal Cancer by a Natural Product 7-Acetylhorminone and Interactions with BSA/HSA: Multispectral Analysis and In Silico and In Vitro Studies

“…The peptide bonds have two orientations of the transition dipole, n−π* and π–π* transitions at 220 and 200 nm for alpha C–O MO transition and alpha N–C MO transitions, respectively, in the secondary region of protein CD spectra. − A high amount of solvent DMF abruptly changes the protein secondary conformations in this region. The DMF effect does not affect the secondary structure of BSA up to 10 μL at 220 and 200 nm.…”

Lung Surfactant Protein B Peptide Mimics Interact with the Human ACE2 Receptor

“…These short amino acid sequences have little secondary conformation, even when bonded in a complex with the receptor protein. The second class of peptide viral fusion blockers have amino acid sequences that span from approximately 20–60 residues in length [ 12 , 16 , 22 ]. These blocking inhibitors have considerable alpha helical propensity and assume a tertiary fold of a helix–hairpin-like structure.…”