Recent findings indicate that nitric oxide (NO*) over-production might be an important factor in the pathogenesis of sporadic amyotrophic lateral sclerosis (SALS). We measured significantly higher concentrations of uric acid and thiol group-containing molecules (R-SH groups) in the cerebrospinal fluid (CSF) from SALS patients compared to controls. The above factors, together with a slightly increased free iron concentration found in the CSF, favour conditions necessary for the formation of the dinitrosyl iron complex, capable of NO* bio-transformation. Thus, we performed ex vivo saturation of CSF (from both SALS patients and controls) with NO*. A decrease in the level of R-SH was found. This was more pronounced in the CSF from SALS patients. In the CSF from SALS patients the production of nitrite and hydroxylamine was greater than that observed in the CSF from controls. Moreover, we also found increased Cu,Zn-SOD activity in the CSF from SALS patients (when compared to control subjects) but no activity corresponding to Mn-SOD in any CSF samples. As Cu,Zn-SOD can react with nitroxyl forming NO*, the conditions for a closed, but continuous, loop of NO* biotransformation are present in the CSF of ALS patients.

In this study we have described the π–π and cation–π interactions between the porphyrin ring and the protein part of porphyrin-containing proteins to better understand their stabilizing role. The number of π–π interactions was higher than that of cation–π interactions in the same set of proteins studied. The pyrrole groups of one porphyrin can be involved in π–π interactions with π systems of another porphyrin in the protein. We have found 5.1% cation–π interactions between porphyrin Fe²⁺ metal cations and π systems of surrounding amino acids as well as the pyrrole rings of other porphyrins. We observed that most of the π–π interactions have an energy in the range −0.5 to −2.0 kcal mol⁻¹, while the cation–π interactions showed an energy in the range −2 to −4 kcal mol⁻¹. Further, an appreciable number of metal/cation–π interaction pairs have an energy in the range −6 to −13 kcal mol⁻¹. The preferred parallel-stacked orientation is found to be more stable than a T-shaped structure for the full set of π–π interaction pairs. In the case of cation–π interactions, it was found that 44% of the cation–π interactions involved planar stacking, 37% of the interactions belonged to the oblique category, and the remaining 19% of the interactions were of the orthogonal type. The separation distance between the cation group and the aromatic ring decreases as the interplanar angle decreases. Furthermore, in the present study we have found that 10.4% of π residues and 3.9% of cationic residues were found to have one or more stabilization centers. Amino acids deployed in the environment of porphyrin rings are deposited in helices and coils. The results from this study might be used for structure-based porphyrin protein prediction and as scaffolds for future porphyrin-containing protein design.