Structure-Based Analysis of Single Nucleotide Variants in the Renin-Angiotensinogen Complex

Abstract

Background: The renin-angiotensin system (RAS) plays an important role in regulating blood pressure and controlling sodium levels in the blood. Hyperactivity of this system has been linked to numerous conditions including hypertension, kidney disease, and congestive heart failure. Three classes of drugs have been developed to inhibit RAS. In this study, we provide a structure-based analysis of the effect of single nucleotide variants (SNVs) on the interaction between renin and angiotensinogen with the aim of revealing important residues and potentially damaging variants for further inhibitor design purposes.

Objectives: To identify SNVs that have functional and potentially damaging effects on the renin-angiotensinogen complex and to use computational approaches to investigate how SNVs might have damaging effects.

Methods: A comprehensive set of all known SNVs in the renin and angiotensinogen proteins was extracted from the HUMA database. This dataset was filtered by removing synonymous and missense variants and using the VAPOR pipeline to predict which variants were likely to be deleterious. Variants in the filtered dataset were modeled into the renin-angiotensinogen complex using MODELLER and subjected to molecular dynamics simulations using GROMACS. The residue interaction networks of the resultant trajectories were analyzed using graph theory.

Conclusions: This research identified important SNVs in the interface of RAS and showed how they might affect the function of the proteins. For instance, the mutant complex containing the variant P40L in angiotensinogen caused instability in the complex, indicating that this mutation plays an important role in disrupting the interaction between renin and angiotensinogen. The mutant complex containing the SNV A188V in renin was shown to have significantly increased fluctuation in the residue interaction networks. D104N in renin, associated with renal tubular dysgenesis, caused increased rigidity in the protein complex comparison to the wild type, which probably in turn negatively affects the function of RAS.

Figure 1

PROSA results for the renin-angiotensinogen complex.

Figure 2

RMSD of the complex containing the SNV, P40L, in angiotensinogen. This is the only mutation tested that destabilizes the complex.

Figure 3

RMSF of ren_D104V. Increased rigidity is noticeable from residue 260 to 380 of renin (chain A) in disease-associated model, ren_D104V.

Figure 4

RMSF ang_P40L. Increased fluctuation is noticeable in ang_P40L between residue 300 and 380 of angiotensinogen (chain B).

Figure 5

A) A heat map depicting the absolute difference between ΔBC in the wild type (first row) and ΔBC in the mutants (ΔΔBC) in chain A (renin) of the complex. Most differences are confined to the color-coded regions. B) The renin-angiotensinogen complex. The areas consisting of large changes in BC in renin (red) have been mapped to the structure.

Figure 6

A) A heat map depicting the absolute difference between ΔBC in the wild type (first row) and ΔBC in the mutants (ΔΔBC) in chain B (angiotensinogen) of the complex. B) The areas consisting of large changes in BC in angiotensinogen (blue) have been mapped to the complex structure.

Figure 7

A) Heat map representing the absolute difference in ΔL/L between the wild type renin and the mutants. The mutant complex, ang_P40L, stands out from residue 245 until the end of renin. B) Closer inspection of ang_P40L shows an elevated ΔL/L for the later part of renin, while the standard deviation, depicted by the red error bars, remains relatively small.

Figure 8

Residue contact map of ang_P40L (left) and the wild type (right) complexes. Interaction with HIS367.A and THR84.A is severely reduced in the mutant complex.

Figure 9

Interface of renin-angiotensinogen wild type. Residues HIS367 and THR84 in renin interact with PRO40 in angiotensinogen. In ang_P40L, Proline is replaced with Leucine and the interaction is severely reduced. In addition, ΔL/L for the region corresponding to residue 245 to 407 (green). As such, it appears that the loss of these two interactions affects the accessibility of this region.

Figure 10

A) A heat map depicting the difference in the standard deviation of the average ΔL/L between wild type and the mutants. The mutant complex, ren_A188V, appears to fluctuate more than the other complexes. B) Large error bars in the ΔL/L plot indicate that ren_A188V fluctuated heavily over the 100ns MD simulation.

Figure 11

The residue contact map for ang_A188V (left) and the wild type (right). The interaction between PHE41.B occurs more often in the mutant complex.