The G2019S pathogenic mutation disrupts sensitivity of leucine‐rich repeat kinase 2 to manganese kinase inhibition

Abstract

J. Neurochem. (2010) 115, 36–46. Abstract Mutations in leucine‐rich repeat kinase‐2 (LRRK2) are the most common cause of late‐onset Parkinson disease. Previously, we showed that the G2019S pathogenic mutation can cause a dramatic increase (approximately 10‐fold) in kinase activity, far above other published studies. A notable experimental difference was the use of Mn‐ATP as a substrate. Therefore, the effects of metal cation‐ATP cofactors on LRRK2 kinase activity were investigated. It is shown, using several divalent metal cations, that only Mg2+ or Mn2+ can support LRRK2 kinase activity. However, for wild‐type, I2020T, and R1441C LRRK2, Mn2+ was significantly less effective at supporting kinase activity. In sharp contrast, both Mn2+ and Mg2+ were effective at supporting the activity of G2019S LRRK2. These divergent effects associated with divalent cation usage and the G2019S mutation were predominantly because of differences in catalytic rates. However, LRRK2 was shown to have much lower (approximately 40‐fold) ATP Km for Mn‐ATP compared with Mg‐ATP. Consequently, sub‐stoichiometric concentrations of Mn2+ can act to inhibit the kinase activity of wild‐type, but not G2019S LRRK2 in the presence of Mg2+. From these findings, a new model is proposed for a possible function of LRRK2 and the consequence of the G2019S LRRK2 pathogenic mutation.

The dual enzyme LRRK2 hydrolyzes GTP in both its GTPase and kinase domains in vitro

“…The efficiency of ATP-mediated kinase activity for WT and G2019S LRRK2 both increased by ~14 and ~8 fold, respectively, compared to Mg 2+ reactions (Figure 2E, F). These results are consistent with previous observations for the effects of Mn 2+ on LRRK2 kinase activity [18, 30]. Therefore, Mn 2+ -bound G2019S LRRK2 showed a large increase (>10-fold) in kinase activity for both autophosphorylation and peptide phosphorylation when either GTP or ATP are used as phospho-donors.…”

“…We therefore substituted Mn 2+ for Mg 2+ and with this increased efficiency we could develop robust GTP-mediated kinase activity. Further, in our experiments using a protein fragment of LRRK2 that lacks the N-terminal domain, we were able to replicate a previously reported finding in full-length LRRK2 protein that G2019S LRRK2 in the Mn 2+ bound state shows a much larger increase in kinase activity relative to WT LRRK2 than Mg 2+ bound LRRK2 [30]. …”

Neurotrophin treatment to promote regeneration after traumatic CNS injury

“…A vital strategy for enhancing recovery from contusive SCI is remyelination. In vitro , NGF and BDNF exhibit a promyelinating influence on DRG explants via activation of neuronally expressed TrkA and oligodendroglial TrkB-FL receptors (Chan et al, 2004; Xiao et al, 2010) respectively. In contrast, in vitro , exogenous NT-3 has been shown to have an inhibitory effect on Schwann cell dependent myelination (Chan et al, 2001; Cosgaya et al, 2002).…”

Down-regulation of LRRK2 in control and DAT transfected HEK cells increases manganese-induced oxidative stress and cell toxicity

“…These observations are particularly relevant to Mn toxicity as Mn can also perturb dopaminergic function (Peneder et al , 2011; Sriram et al , 2010) and inhibit amphetamine-induced DA release (Guilarte et al , 2008) as well as promote mitochondria degeneration. Interestingly, Mn can increase the activity of the most prevalent mutant form of LRRK2, G2019S, but inhibit wild-type kinase activity which is normally stimulated by Mg (Covy and Giasson, 2010; Lovitt et al , 2010). Thus, LRRK2 displays a number of critical functions many of which can affect or be affected by Mn.…”