Ab initio structures and stabilities of helide cations: HeX

The electronic structure and molecular properties of helides of the form ${\rm HeX}^{n+}$ (where $\rm X=B{-}Ne$, Al-Ar and $n=1{-}3$) were studied using the CCSD(T) method in conjunction with the series of correlation-consistent basis sets. The highest level of theory employed, the CCSD(T)-FC/cc-pVQZ model, was used to elucidate trends in bond lengths, dissociation energies and harmonic frequencies. The more highly charged species were found to have shorter bond lengths than the singly charged species. The ground states of the helide cations were often those with longer bond lengths when compared with the excited state ions

An Eckart-Watson Hamiltonian for linear molecules in the rectilinear displacement w-coordinates and an application to HCN

A 3N-5 Eckart-Watson vibration-rotation Hamiltonian for linear triatomic molecules was developed in terms of rectilinear displacement w-coordinates. The w-coordinate system has been developed in order to minimize vibration- rotation coupling. For numerical simplicity, a 3N-6 Hamiltonian was obtained by constraining the bending vibrations to the xz-plane. Vibrational (J = 0) solution algorithms for both four- and three-dimensional representations utilize finite-element method and grid techniques (Gaussian quadrature and discrete variable representation). The validities and accuracies of the Hamiltonians and their solution algorithms have been examined using a recent ab initio potential energy surface [A.T. Wong, G.B. Bacskay, Mol. Phys. 79 (1993) 819] to calculate the vibrational eigenenergies of the HCN molecule

Characterisation of tryptic peptides of phosphorylated tyrosine hydroxylase by high-pressure liquid chromatography electrospray ionisation mass spectrometry

Tyrosine hydroxylase (TH) is involved in the biosynthesis of catecholamines and is activated by phosphorylation. Phosphorylated TH was analysed using high-pressure liquid chromatogaphy combined with electrospray mass spectrometry (HPLC ESI-MS). Two mass scanning methods were used to detect tryptic cleavage products of TH. In the positive electrospray ionisation mode (ESI+), the peptides that contain the phosphorylation sites of TH were identified. In the alternative method, a phosphopeptide was detected in the negative electrospray ionisation mode (ESI-) using single ion monitoring in combination with a sequential ESI+ switching experiment. A raised baseline interfered with detection of hydrophilic peptides in ESI-, with the signal-to-noise ratio indicating that the method was operating near the limit of detection for a conventional electrospray source. The switching method improved the certainly of identification of phosphopeptides

Tyrosine hydroxylase phosphorylation: regulation and consequences

The rate-limiting enzyme in catecholamine synthesis is tyrosine hydroxylase. It is phosphorylated at serine (Ser) residues Ser8, Ser19, Ser31 and Ser40 in vitro, in situ and in vivo. A range of protein kinases and protein phosphatases are able to phosphorylate or dephosphorylate these sites in vitro. Some of these enzymes are able to regulate tyrosine hydroxylase phosphorylation in situ and in vivo but the identity of the kinases and phosphatases is incomplete, especially for physiologically relevant stimuli. The stoichiometry of tyrosine hydroxylase phosphorylation in situ and in vivo is low. The phosphorylation of tyrosine hydroxylase at Ser40 increases the enzyme's activity in vitro, in situ and in vivo. Phosphorylation at Ser31 also increases the activity but to a much lesser extent than for Ser40 phosphorylation. The phosphorylation of tyrosine hydroxylase at Ser19 or Ser8 has no direct effect on tyrosine hydroxylase activity. Hierarchical phosphorylation of tyrosine hydroxylase occurs both in vitro and in situ, whereby the phosphorylation at Ser19 increases the rate of Ser40 phosphorylation leading to an increase in enzyme activity. Hierarchical phosphorylation depends on the state of the substrate providing a novel form of control of tyrosine hydroxylase activation

Phosphorylation of Ser¹⁹ alters the conformation of tyrosine hydroxylase to increase the rate of phosphorylation of Ser⁴⁰

The effect of phosphorylation on the shape of tyrosine hydroxylase (TH) was studied directly using gel filtration and indirectly using electrospray ionization mass spectrometry. Phosphorylation of Ser¹⁹ and Ser⁴⁰ produced a TH molecule with a more open conformation than the non-phosphorylated form. The conformational effect of Ser¹⁹ phosphorylation is less pronounced than that of the Ser⁴⁰ phosphorylation. The effect of Ser¹⁹ and Ser⁴⁰ phosphorylation appears to be additive. Binding of dopamine produced a more compact form when compared with the non-dopamine-bound TH. The interdependence of Ser¹⁹ and Ser⁴⁰ phosphorylation was probed using electrospray ionization mass spectrometry. The rate constants for the phosphorylation of Ser¹⁹ and Ser⁴⁰ were determined by electrospray ionization mass spectrometry using a consecutive reaction model. The rate constant for the phosphorylation of Ser⁴⁰ is ~2- to 3-fold higher if Ser¹⁹ is already phosphorylated. These results suggest that phosphorylation of Ser¹⁹ alters the conformation of tyrosine hydroxylase to allow increased accessibility of Ser⁴⁰ to kinases