Measuring the Reversible Oxidation of Protein Tyrosine Phosphatases Using a Modified Cysteinyl-Labeling Assay.

The modified cysteinyl-labeling assay enables the labeling, enrichment, and detection of all members of the protein tyrosine phosphatase (PTP) superfamily that become reversibly oxidized in cells to facilitate phosphorylation-dependent signaling. In this chapter, we describe the method in detail and highlight the pitfalls of avoiding post-lysis oxidation of PTPs to measure the dynamic and transient oxidation and reduction of PTPs in cell signaling.

Protein tyrosine phosphatase 1B regulates miR-208b-argonaute 2 association and thyroid hormone responsiveness in cardiac hypertrophy.

Increased production of reactive oxygen species plays an essential role in the pathogenesis of several diseases, including cardiac hypertrophy. In our search to identify redox-sensitive targets that contribute to redox signaling, we found that protein tyrosine phosphatase 1B (PTP1B) was reversibly oxidized and inactivated in hearts undergoing hypertrophy. Cardiomyocyte-specific deletion of PTP1B in mice (PTP1B cKO mice) caused a hypertrophic phenotype that was exacerbated by pressure overload.

In Vitro Activity Assays to Quantitatively Assess the Endogenous Reversible Oxidation State of Protein Tyrosine Phosphatases in Cells.

The reversible oxidation of protein tyrosine phosphatases (PTPs) impairs their ability to dephosphorylate substrates in vivo. This transient inactivation of PTPs occurs as their conserved catalytic cysteine residue reacts with cellular oxidants thereby abolishing the ability of this reactive cysteine to attack the phosphate of the target substrate. Hence, in vivo, the inhibition of specific PTPs in response to regulated and localized rises in cellular oxidants enables phospho-dependent signaling.

Regulation of PTP1B activation through disruption of redox-complex formation.

We have identified a molecular interaction between the reversibly oxidized form of protein tyrosine phosphatase 1B (PTP1B) and 14-3-3ζ that regulates PTP1B activity. Destabilizing the transient interaction between 14-3-3ζ and PTP1B prevented PTP1B inactivation by reactive oxygen species and decreased epidermal growth factor receptor phosphorylation. Our data suggest that destabilizing the interaction between 14-3-3ζ and the reversibly oxidized and inactive form of PTP1B may establish a path to PTP1B activation in cells.

Expression, purification and crystallization of acetyl-CoA hydrolase from Neisseria meningitidis.

Neisseria meningitidis is the causative microorganism of many human diseases, including bacterial meningitis; together with Streptococcus pneumoniae, it accounts for approximately 80% of bacterial meningitis infections. The emergence of antibiotic-resistant strains of N. meningitidis has created a strong urgency for the development of new therapeutics, and the high-resolution structural elucidation of enzymes involved in cell metabolism represents a platform for drug development.

Balloon atrial septostomy through internal jugular vein in a 45-day-old child with transposition of great arteries.

Balloon atrial septostomy is a common palliative procedure in D-transposition of great arteries. It is technically easy before 2-3 weeks of age when the septum primum is thin. Femoral vein or umbilical vein, when available, is the common access used for this procedure.

Veno-venous shunt-assisted cavopulmonary anastomosis.

Objective: The bidirectional Glenn shunt is commonly performed under cardiopulmonary bypass for conditions that lead to a single ventricle repair. We report our experience of bidirectional Glenn shunt done without cardiopulmonary bypass.

Methods: Between June 2007 and May 2009, 186 consecutive patients underwent off-pump bidirectional Glenn shunt for a variety of complex cyanotic congenital heart defects.

Tricuspid valve repair with neopapillary muscle from right ventricular trabecula.

Congenital abnormalities of the tricuspid valve without downward displacement of the septal and posterior tricuspid leaflets are uncommon causes of tricuspid regurgitation. Progressive tricuspid regurgitation can lead to right heart dilatation, arrhythmia, and irreversible deterioration of right ventricular function. Tricuspid valve repair is an important method to stabilize symptomatic children because tricuspid valve replacement has a poor prognosis.