Balancing reactivity against selectivity: the evolution of protein S-nitrosylation as an effector of cell signaling by nitric oxide

Abstract

Produced by the action of lightning in the atmosphere of the pre-biotic earth, nitric oxide (NO) is a free radical molecule that provided the major nitrogen source for development of life. Remarkably, when atmospheric sources of NO became restrictive, organisms evolved the capacity for NO biosynthesis and NO took on bioregulatory roles. We now recognize NO as an ancestral regulator of diverse and important biological functions, acting throughout the phylogenetic tree. In mammals, NO has been implicated as a pivotal regulator of virtually every major physiological system. The bioactivities of NO, and reactive species derived from NO, arise predominantly from their covalent addition to proteins. Importantly, S-nitrosylation of protein cysteine (Cys) residues has emerged as a preeminent effector of NO bioactivity. How and why NO selectively adds to particular Cys residues in proteins is poorly understood, yet fundamental to how NO communicates its bioactivities. Also, evolutionary pressures that have shaped S-nitrosylation as a biosignaling modality are obscure. Considering recently recognized NO signaling paradigms, we speculate on the origin of NO signaling in biological systems and the molecular adaptations that have endowed NO with the ability to selectively target a subset of protein Cys residues that mediate biosignaling.

Figure 1

Homology model of human argininosuccinate synthase (AS). A: Alignment of some amino acid sequences of human AS and orthologs, showing Cys132 and phylogenetic substitutions. The depicted result and those extended to 130 additional orthologs, suggest that a Cys132 homolog is present in NOS-expressing species, but absent in non-NOS expressing species. This is in accord with the view that Cys132 evolved as a specific NO-sensor. B: Model structure of AS monomer. The model reveals that Cys132 of human AS resides in the substrate-binding helix, shown in red. Three substrates, ATP, Asp and Cit are shown in pink, cyan and yellow respectively. C: Close up view of the AS active site, showing bound substrates and the Cys132 environment. Sulfur of Cys 132 is depicted as a yellow sphere, and substrates are rendered in ball and stick form, with ATP in black and Asp and Cit in gray. Also shown are hydrophobic residues (pink) surrounding Cys132 (A99, V103, V114, L133, I139). Residues involved in hydrogen-bonding of substrates are indicated (N123, gray; D124, green; Q125, gray; R127, blue) as well as aromatic residues in proximity to Cys132 (F128 and Y133, orange). Residues potentially involved in acid-base catalysis, H116 and E129, are shown in cyan and green respectively. Results suggest that Cys132 is locating on a conserved substrate-binding α-helix. S-nitrosylation of Cys132 would destabilize the helix and have adverse consequences on either binding of substrates or positioning substrates for productive catalysis.