NOSs

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Introduction

Nitric oxide (NO) is a messenger molecule with numerous molecular targets. It can regulate neurotransmission or vascular tone, gene transcription, mRNA translation, and produce post-translational modifications of proteins. The reaction of NO and superoxide anion (O₂^-) to form the potent oxidant peroxynitrite (ONOO^-) is an important way for NO inactivation. ONOO^- can cause oxidative damage, nitration, and S-nitrosylation of biomolecules including proteins, lipids and DNA.

In mammals, NO can be produced by three different isoenzymes of NO synthase (NOS; L-arginine, NADPH: oxygen oxidoreductases, NO forming; EC 1.14.13.39). These three isozymes are called neuronal 'n' NOS (or NOS I), inducible 'i' NOS (or NOS II) and endothelial 'e' NOS (or NOS III), respectively (Figure 1).

Figure 1. Important functions of the different NOS isoforms (Förstermann, U.; Sessa, W.C. 2012)

Mechanisms of nitric oxide synthesis

All NOS proteins are homodimers (Figure 2). L-arginine is a substrate used by all isoforms of NOS, and molecular oxygen and reduced nicotinamide adenine dinucleotide phosphate (NADPH) are used as co-substrates. The cofactors of all isoenzymes are flavin adenine dinucleotide (FAD), flavin mononucleotide (FMN) and (6 R-)5,6,7,8-tetrahydro-L-biopterin (BH 4). A functional NOS transfers electrons from NADPH to the haem in the amino-terminal oxygenase domain. The electrons in the haem site are responsible for reducing and activating O₂ and oxidizing L-arginine to L-citrulline and NO.

The NOS enzyme goes through two steps to synthesize NO. First, NOS hydroxylates L-arginine to N^ω-hydroxy-L-arginine. NOS then oxidizes N^ω-hydroxy-L-arginine to L-citrulline and NO. All isoforms of NOS bind calmodulin. In nNOS and eNOS, calmodulin binding is brought about by an increase in intracellular Ca²⁺. In iNOS, due to the different amino acid structure of the calmodulin binding site, calmodulin binds under very low Ca²⁺ concentration (less than 40 nM). All NOS proteins contain a zinc-thiolate cluster formed by zinc ions, where zinc has a structural function rather than a catalytic function.

Figure 2. Structure and catalytic mechanisms of functional NOS (Förstermann, U.; Sessa, W.C. 2012)

Neuronal nitric oxide synthase

Neuron NOS is expressed in specific neurons of the brain. The nNOS in the brain exists in particulate and soluble forms, and the localization of its different sub-cells helps it achieve diversified functions. Neuronal NOS contains a PDZ domain, which can directly interact with the PDZ domains of other proteins. These important interactions determine the subcellular distribution and activity of enzymes. In addition to brain tissue, nNOS has also been identified in the spinal cord, sympathetic ganglia, adrenal glands, peripheral nitrergic nerves, epithelial cells of various organs, and pancreatic islet cells, etc. In mammals, skeletal muscle is the largest source of nNOS.

Conclusion

All three NOS isoenzymes play important regulatory functions in the cardiovascular system. Neuronal NOS is involved in the central regulation of blood pressure, and nerves containing nNOS can dilate certain vascular beds. Phosphodiesterase 5 inhibitors require at least residual nNOS activity to perform their effects. Studies have found that inducible NOS is expressed in atherosclerotic plaques, indicating that it is an important mediator of the fall in blood pressure in septic shock. eNOS has very important functions, for example, it can keep blood vessel dilation, control blood pressure, and has many other vasoprotective and anti-atherosclerosis effects. Although there is no research to prove that eNOS is a "disease gene", many cardiovascular risk factors can cause eNOS uncoupling, oxidative stress, and vascular endothelial dysfunction. Drugs that interfere with the renin-angiotensin-aldosterone system and statins can be used to prevent endothelial dysfunction. Subsequent research should further clarify how these therapeutic agents promote eNOS coupling when oxidative stress increases, so as to discover and gain insight into other potential pathways that lead to a beneficial effect of NO in the cardiovascular system.