From oncoproteins to spike proteins: the evaluation of intramolecular stability using hydropathic force field

Abstract

Evaluation of the intramolecular stability of proteins plays a key role in the comprehension of their biological behavior and mechanism of action. Small structural alterations such as mutations induced by single nucleotide polymorphism can impact biological activity and pharmacological modulation. Covid-19 mutations, that affect viral replication and the susceptibility to antibody neutralization, and the action of antiviral drugs, are just one example. In this work, the intramolecular stability of mutated proteins, like Spike glycoprotein and its complexes with the human target, is evaluated through hydropathic intramolecular energy scoring originally conceived by Abraham and Kellogg based on the "Extension of the fragment method to calculate amino acid zwitterion and side-chain partition coefficients" by Abraham and Leo in Proteins: Struct. Funct. Genet. 1987, 2:130 - 52. HINT is proposed as a fast and reliable tool for the stability evaluation of any mutated system. This work has been written in honor of Prof. Donald J. Abraham (1936-2021).

Keywords: Intramolecular stability; LogPo/w; Mutations; Scoring function.

Fig. 1

 K and N-RAS Intramolecular HINT score. K-RAS: The intramolecular stabilities of the mutated inactive (PDB ID: 5W22) and active (PDB ID: 6GOD) conformations of K-RAS were calculated. G12C, G12D, G12V, G13D, G13C, and Q61H are the most stable K-RAS closed conformations and present the highest experimental frequency. H-RAS: The intramolecular stability of the mutated inactive (PDB ID: 6WGH) and active (PDB ID: 5UHV) conformations reveal that G12D, G12S, and Q61R are the most stable and frequent N-RAS closed conformations [16].

Fig. 2

H-RAS structures and Intramolecular HINT score. Three different conformations of H-RAS were identified: an inactive GDP-bound conformation (blue) (PDB ID: 4Q21), an active state 1 conformation (orange) (PDB ID: 3RSO), and an active state 2 conformation (grey) (PDB ID: 5P21). The ΔHINT score between the mutated and the wild-type structure was calculated considering all three possible conformations. Only state 2 active conformation is stabilized by mutations

Fig. 3

B-RAF Intramolecular HINT scores. The ΔHINT score between mutated and wildtype structure (PDB ID: 6XFP) was calculated. All mutated forms are more stable than the wild-type ones. V599D is the most stable and presents the higher experimental kinase activity

Fig. 4

Intramolecular HINT output file. The output file shows each atom-atom interaction and the total intramolecular score with a detailed description of the single energy contribution (hydrogen bond, electrostatic and hydrophobic interactions, and negative contributions). In this capture, for example, the mutated ARG12 establishes two different hydrogen bonds with the residue GLU62.

Fig. 5

Intramolecular HINT score of Spike proteins. All mutants generated from the trimeric wildtype closed conformation (PDB ID: 6VXX) are as stable as the wild type. RBD HINT score reveals higher stability of mutants than wildtype. Alpha and Omicron variants present a higher affinity toward ACE2, and their complexes are the most stable