Determination of the endogenous phosphorylation state of B‐50/GAP‐43 and neurogranin in different brain regions by electrospray mass spectrometry

Neurogranin, a link between calcium/calmodulin and protein kinase C signaling in synaptic plasticity

“…However, to my knowledge, in vivo acylation of Ng cysteines has not been reported to date. The mass-spectrometry data obtained in rat brain Ng (60,61) and the high Ng solubility when extracted from brain homogenates do not support the existence of such modifications in vivo. In a previous study, we showed that Ng binds to PA and that the intracellular distribution of Ng is affected by overexpression of PLD, an enzyme that catalyzes the production of PA from phosphatidylcholine (PLC) (76).…”

“…Several studies have analyzed the in vivo phosphorylation status of rat brain Ng using mass spectrometry. While initial work proposed that phospho-Ng is the major isoform in vivo (60), later studies questioned these data and showed that the N-terminus was acetylated in vivo (61). Our results, using phospho-Ng specific antibodies, suggest the presence of low amounts of phosphorylated Ng in freshly extracted rat brain.…”

Determination of Phosphorylation Levels of Tyrosine Hydroxylase by Electrospray Mass Spectrometry

“…This will be of particular relevance to work carried out in vivo (21) to determine the endogenous phosphorylation levels of TH. Application of this procedure to species other than rat is possible because the amino acid sequence of bovine and human TH contains similar cleavage points to produce potential TH-P/TH markers.…”

“…The underlying rationale for using protein fragments as markers is to ensure that unit mass resolution is obtained for the fragment peak. They extended this approach using ESMS to determine endogenous phosphorylation of B50/GAP-43 and neurogranin in different regions of the brain (21).…”

Microscale Purification of Proteins Exhibiting Anomalous Electrophoretic Migration: Application to the Analysis of GAP-43 Phosphorylation

“…It is well known that GAP-43 is phosphorylated by PKC at serine 41 and that this phosphorylation prevents its interaction with calmodulin (3,13). More recently, additional phosphorylation sites have been reported (14,15), but their physiological significance is not known.…”