Effect of dodecylsulfate binding on the activity of creatine kinase

The number of lithium dodecylsulfate (LDS) molecules tightly bound to creatine kinase has been found by isotachophoresis to be 87 at 25°C upon saturation of the enzyme dimer with LDS. The binding shows positive cooperativity by both the Hill and Scatchard plots. The enzyme is completely inactivated when its high-affinity sites are fully occupied with LDS. However, at partial LDS saturation the activity remaining is definitely higher than can be accounted for by the amount of free enzyme left, showing the presence of species of active molecules with the tight LDS sites only partly saturated. The presence of ATP leads to a decrease in detergent bound at the high-affinity sites with partial restoration of activity.     

Nitric oxide suppresses LPS-induced inflammation in a mouse asthma model by attenuating the interaction of IKK and Hsp90

A feature of allergic airway disease is the observed increase of nitric oxide (NO) in exhaled breath. Gram-negative bacterial infections have also been linked with asthma exacerbations. However, the role of NO in asthma exacerbations with gram-negative bacterial infections is still unclear. In this study, we examined the role of NO in lipopolysaccharide (LPS)-induced inflammation in an ovalbumin (OVA)-challenged mouse asthma model. To determine whether NO affected the LPS-induced response, a NO donor (S-nitroso-N-acetylpenicillamine, SNAP) or a selective inhibitor of NO synthase (1400W) was injected intraperitoneally into the mice before the LPS stimulation. Decreased levels of proinflammatory cytokines were demonstrated in the bronchoalveolar lavage fluid from mice treated with SNAP, whereas increased levels of cytokines were found in the 1400W-treated mice. To further explore the molecular mechanism of NO-mediated inhibition of proinflammatory responses in macrophages, RAW 264.7 cells were treated with 1400W or SNAP before LPS stimulation. LPS-induced inflammation in the cells was attenuated by the presence of NO. The LPS-induced IκB kinase (IKK) activation and the expression of IKK were reduced by NO through attenuation of the interaction between Hsp90 and IKK in the cells. The IKK decrease in the lung immunohistopathology was verified in SNAP-treated asthma mice, whereas IKK increased in the 1400W-treated group. We report for the first time that NO attenuates the interaction between Hsp90 and IKK, decreasing the stability of IKK and causing the down-regulation of the proinflammatory response. Furthermore, the results suggest that NO may repress LPS-stimulated innate immunity to promote pulmonary bacterial infection in asthma patients.