טכניון מכון טכנולוגי לישראל
הטכניון מכון טכנולוגי לישראל - בית הספר ללימודי מוסמכים  
M.Sc Thesis
M.Sc StudentAbu Hariri Hiba
SubjectThe Role of Trx-related Protein of 14kDa(TRP14)
in Regulating Protein S-Nitrosylation
DepartmentDepartment of Medicine
Supervisor Professor Moran Benhar
Full Thesis textFull thesis text - English Version


Abstract

S-Nitrosylation, the covalent addition of a nitroso group to a cysteine thiol to form S-nitrosothiol (SNO), has been shown to modify the function of a broad spectrum of proteins and thereby to convey the ubiquitous influence of nitric oxide (NO) on signal transduction and cellular responses. Elevated nitrosylation that occurs during nitrosative stress has been implicated in cell dysfunction and pathological processes. The redox protein thioredoxin 1 (Trx1) has been demonstrated to denitrosylate various proteins and thereby to regulate NO/nitrosylation-mediated cellular processes and to protect cells from nitrosative stress. Trx-related protein of 14 kDa (TRP14), a recently identified member of the Trx family, is a ubiquitous cytosolic protein whose cellular role is not well understood. It shares 20% sequence identity and 37% similarity with human Trx1 and exhibits a typical Trx fold but also structural features that differ from Trx1. Despite their sequence and structure similarities, TRP14 lacks activity with the classical disulfide substrates of Trx1. Recent evidence suggests that TRP14 acts as an endogenous denitrosylase.

In this study, we investigated the role of TRP14 in regulating protein S-nitrosylation. As a first step, we showed that similar to Trx, TRP14 can catalyze the reduction of low molecular weight SNO, such as S-nitrosoglutathione. In addition, using biochemical and redox proteomics approaches, we found that TRP14 catalyzes the denitrosylation of multiple cellular proteins, albeit less efficiently than Trx1. The proteomic analysis revealed a subset of distinct SNO targets of TRP14 that differ from Trx1, but also shared substrates. Finally, in a cell free assay, we found that, unlike Trx1, TRP14 is resistant to inhibition by high concentrations of NO. Our findings suggest that TRP14, by its denitrosylase function, could be an important element in the regulation NO/SNO-dependent cellular responses and may play a role in the cellular response to nitrosative stress.