ABSTRACT
Translationally controlled tumor protein (TCTP) is a potential target for cancer therapy. It functions as a growth regulating protein implicated in the TSC1-TSC2 -mTOR pathway or a guanine nucleotide dissociation inhibitor for the elongation factors EF1A and EF1Bbeta. Accumulating evidence indicates that TCTP also functions as an antiapoptotic protein, through a hitherto unknown mechanism. In keeping with this, we show here that loss of tctp expression in mice leads to increased spontaneous apoptosis during embryogenesis and causes lethality between E6.5 and E9.5. To gain further mechanistic insights into this apoptotic function, we solved and refined the crystal structure of human TCTP at 2.0 A resolution. We found a structural similarity between the H2-H3 helices of TCTP and the H5-H6 helices of Bax, which have been previously implicated in regulating the mitochondrial membrane permeability during apoptosis. By site-directed mutagenesis we establish the relevance of the H2-H3 helices in TCTP's antiapoptotic function. Finally, we show that TCTP antagonizes apoptosis by inserting into the mitochondrial membrane and inhibiting Bax dimerization. Together, these data therefore further confirm the antiapoptotic role of TCTP in vivo and provide new mechanistic insights into this key function of TCTP.
Subject(s)
Apoptosis , Biomarkers, Tumor/chemistry , Biomarkers, Tumor/metabolism , Mitochondria/metabolism , bcl-2-Associated X Protein/metabolism , Amino Acid Sequence , Animals , Biomarkers, Tumor/genetics , Cell Line , Crystallography, X-Ray , Dimerization , Embryonic Development , Humans , Mice , Mice, Knockout , Molecular Sequence Data , Mutation , Protein Structure, Secondary , Protein Structure, Tertiary , Recombinant Proteins/chemistry , Recombinant Proteins/metabolism , Sequence Alignment , Tumor Protein, Translationally-Controlled 1 , bcl-2-Associated X Protein/chemistryABSTRACT
This paper describes the developments, role and contributions of the NMR spectroscopy groups in the Structural Proteomics In Europe (SPINE) consortium. Focusing on the development of high-throughput (HTP) pipelines for NMR structure determinations of proteins, all aspects from sample preparation, data acquisition, data processing, data analysis to structure determination have been improved with respect to sensitivity, automation, speed, robustness and validation. Specific highlights are protonless (13)C-direct detection methods and inferential structure determinations (ISD). In addition to technological improvements, these methods have been applied to deliver over 60 NMR structures of proteins, among which are five that failed to crystallize. The inclusion of NMR spectroscopy in structural proteomics pipelines improves the success rate for protein structure determinations.
Subject(s)
Magnetic Resonance Spectroscopy/methods , Proteomics/methods , Algorithms , Data Interpretation, Statistical , Models, Molecular , Proteins/chemistryABSTRACT
A 29 year old patient with alpha-1 antitrypsin deficiency and bullous emphysema became pregnant against the advice of her physicians. Despite a mid-trimester pneumothorax requiring the insertion of a chest tube, she went on to deliver a healthy child under epidural anaesthesia using a midforceps technique. Vaginal delivery is not necessarily contra-indicated in multiparous patients with bullous emphysema.
Subject(s)
Pregnancy Complications , Pulmonary Emphysema/etiology , alpha 1-Antitrypsin Deficiency , Adult , Female , Humans , Infant, Newborn , Labor, Obstetric , Male , PregnancyABSTRACT
In order to assess the incidence of Legionnaires' disease in a district general hospital, 90 consecutive patients with pneumonia seen over a 3-year period were investigated. Only 2 patients were found to have Legionnaires' disease, indicating that it is not a frequent cause of pneumonia in the Bristol area.
Subject(s)
Legionnaires' Disease/epidemiology , Adolescent , Adult , Aged , England , Female , Humans , Male , Middle AgedABSTRACT
Phosphoenolpyruvate carbosylase (EC 4.1.1.31) has been used in developing a simple, inexpensive colorimetric assay for serum total carbon dioxide in an open system. The oxaloacetate formed by the action of the enzyme on bicarbonate and phosphoenolpyruvate is measured by use of the diazonium salt of Fast Violet B. Interferences from billirubin, pyruvate, and drugs are negligible. Acetoacetate interference is significant only in highly ketotic samples, and a serum blank corrects for it. Serum protein interference is equivalent to 3.3 plus or minus 1.25 mmol of CO2 per liter and hence is sufficiently constant to be corrected for by use of a serum standard or serum blank. The method has been applied to the Vickers M-300 and D-300 systems and within-batch standard deviations of plus or minus 0.1 to plus or minus 0.6 mmol/liter have been observed. Excellent correlation with orthodox techniques has been obtained.
