266 research outputs found
Structural and dynamic properties of translocase motor SecA:
No full textSecA is a large, 204 kDa, homodimeric, helicase-like protein that is a key component of the bacteria protein secretion machinery. SecA, being a motor protein, couples the translocation of polypeptide segments across or into biological membranes with the expenditure of metabolic energy extracted from ATP hydrolysis. SecA adopts a compact conformation in the cytoplasm but switches to a relaxed one when is engaged to translocation at the membrane. Specific interaction of SecA with SecYEG induces large conformational changes to both partners that result in the stimulation of SecA’s ATPase activity and trigger the opening of the channel.
We use a combination of NMR spectroscopy, Isothermal Titration Calorimetry (ITC) and biochemical techniques to characterize E. coli SecA along the protein secretion pathway. Recent advances in isotope labeling and NMR methodology (methyl-TROSY) enabled the NMR study of SecA.
We found that the nucleotide binding cleft of SecA exists in a metastable state that undergoes a disorder-order transition upon nucleotide binding. Our data show that SecA uses a novel mechanism wherein conserved regions lining the cleft undergo cycles of disorder-order transitions while switching among functional catalytic states. The structural relation of SecA to helicases suggests that these proteins may utilize similar mechanisms to convert the ATP binding/hydrolysis energy to mechanical work.
Our data reveal that C domain undergoes cycles of detachment and rebinding to the motor that are linked with the ATPase activation of SecA. When the contacts with the C domain are loosened, SecA becomes activated and this process is probably facilitated by the membrane. Moreover, allosteric communication between the preprotein binding and the motor domain of SecA is regulated by nucleotide and signal peptide binding. Finally, we found that the extreme C terminus of SecY (C6 loop) contacts both the motor and the C domain of SecA and we identified important residues from the loop C6 that mediate this interaction.
In summary, our studies revealed the intimate relation between flexibility and catalytic efficiency in SecA as well the allosteric communication among the domains.Ph.D.Includes bibliographical references (p. 131-144)by Dimitra Keramisano
Geolinguistic spaces and the geopolitics of organization studies: movements across centers and peripheries: European group for organisational studies colloquium
No full textAcademic knowledge production and evaluation has been studied as a phenomenon situated across global and (semi)peripheral contexts (Lillis and Curry, 2010; Bennett, 2014). The metaphor of scales draws attention to the study of academic discourses as practices that develop across vertical, hierarchically organized layers (Blommaert, 2010). In this paper, we draw on 3 sub-corpora of organization studies journal articles, with each journal occupying a different position in the impact factor continuum. Building on the work of Lillis and Curry (2010), we study networked activities as social capital in the global academic economy. In our analysis we first examine the epistemological profile of organisation studies, as reflected in differentially ranked journals located in European contexts. We then explore the complex configuration of spaces/localities within and around which the production of knowledge takes place, by examining networks of author affiliations in relation to the identified sites in the empirical papers included in the corpus. Finally, we examine how constructions of the ‘local’ and the ‘global’ relate to the themes foregrounded in the aims and scope of each journal and we explore how discourses of the ‘local’ and the ‘global’ relate to key concepts in the field of organization studies, including institutions, alternatives and resistance, as well as power and gender
Immunocytochemical detection of apoptotic markers in smears from precancerous and cancerous lesions of the uterine cervix: correlation with HPV infection and cytomorphologic findings in conventional and ThinPrep cervical samples
No full textIn the present study, cervical smears from women with intraepithelial lesionsand with normal cytology were analyzed morphologically and with the use of immunocytochemical methods. The aim of our morphologic study was to reevaluate the reproducibility of the automated processing of the cervical samples collected with the ThinPrep (TP) method, and to further investigate the potential diagnostic value of additional slides prepared from residual TP Pap Tests. Our results suggest that repeat processing of residual cervical TP samples may not be an invariably reproducible procedure and the first slide may not be necessarily representative of the specimen as a whole. An additional finding of our study is that an upgrading of the cytological diagnosis after repeat processing is more likely to occur in cases initially classified as “ASC-US” or as “squamous intraepitheliallesion, grade cannot be determined” than in cases with an initial negative or HSIL diagnosis. The aim of our immunocytochemical study was to detect the expression pattern of p53, PTEN, Fas, p16 and L1 proteins in intraepithelial lesions and to investigate their potential correlations to each other as well as their value asprognostic markers in LSIL cases. Our results suggest the following : a) loss of PTEN or Fas expression and p53 overexpression may be independently involved, even as early events, in the process of neoplastic transformation of the cervical epithelium by facilitating transition to higher grade dysplasia b)a strong nuclear and cytoplasmic immunocytochemical expression pattern ofp16 might be a strong indicator for the presence of a high grade lesion, and could serve as a useful diagnostic adjunct especially in the presence of equivocal cytomorphologic findings and c) the combined immunocytochemical expression pattern of p16 and L1 proteins may also confer significant diagnostic information as regards the determination of lesion grade and probably contribute in the identification of women at greater risk for developing invasive carcinoma, even when the HPV status is unknown. The potential prognostic value of our examined markers in LSIL cases was not verified in our study and should be further evaluated in future large prospective series.Στην παρούσα μελέτη μελετήθηκαν τόσο μορφολογικά όσο και με ανοσοκυτταροχημικές μεθόδους τραχηλικά επιχρίσματα κατά Παπανικολάου από γυναίκες με ενδοεπιθηλιακές αλλοιώσεις του τραχήλου της μήτρας και με φυσιολογικά ευρήματα. Σκοπός της μορφολογικής μας μελέτης ήταν η επαναξιολόγηση της αναπαραγωγιμότητας της τεχνικής επεξεργασίας των τραχηλικών δειγμάτων υγρής φάσης ThinPrep καθώς και της πιθανής διαγνωστικής αξίας της μεθόδου παρασκευής επιπρόσθετων πλακιδίων εκ του υπολειπόμενου υλικού. Τα αποτελέσματά μας έδειξαν ότι η επαναλαμβανόμενη επεξεργασία των τραχηλικών δειγμάτων ΤΡ μπορεί να μην είναι πάντα και ανεξαιρέτως μια αναπαραγώγιμη διαδικασία και το αρχικό πλακίδιο που παρασκευάζεται μπορεί να μην είναι σε κάθε περίπτωση αντιπροσωπευτικό του συνόλου του δείγματος.Η μελέτη μας κατέδειξε επίσης ότι η πιθανότητα αναθεώρησης της αρχικής διάγνωσης μετά την αξιολόγηση των επαναληπτικών πλακιδίων ThinPrep είναι σημαντικά μεγαλύτερη σε αμφιλεγόμενα περιστατικά, όπως οι περιπτώσεις ASCUS(atypical squamous cells of undetermined significance/ άτυπα πλακώδηκύτταρα αδιευκρίνιστης σημασίας) ή οι δύσκολες διαγνωστικά περιπτώσεις με στοιχεία χαμηλού βαθμού δυσπλασίας στις οποίες δεν μπορεί με βεβαιότητα να αποκλειστεί και η πιθανότητα υψηλόβαθμης αλλοίωσης.Σκοπός της ανοσοκυτταροχημικής μας μελέτης ήταν η διερεύνηση της συχνότητας και του προτύπου πιθανής διαταραχής της έκφρασης των πρωτεϊνώνp53, PTEN, Fas, p16 και L1 στις ενδοεπιθηλιακές αλλοιώσεις του τραχήλου της μήτρας, των πιθανών αλληλοεπιδράσεων των δεικτών αυτών μεταξύ τους καθώς και της πιθανής αξίας της ανοσοκυτταροχημικής τους έκφρασης ως δεικτών πρόγνωσης στις χαμηλού βαθμού δυσπλασίες. Τα αποτελέσματά μας έδειξαν : α)ότι η διαταραχή της έκφρασης των πρωτεϊνών p53, PTEN, και Fas μπορεί να παρατηρηθεί ακόμη και στο ενδοεπιθηλιακό στάδιο της καρκινογένεσης στον τράχηλο της μήτρας, και ενδέχεται να διευκολύνει τη μετάβαση από τη χαμηλού βαθμού στην υψηλού βαθμού δυσπλασία β)ότι η ανοσοκυτταροχημική κατάδειξη της υπερέκφρασης της πρωτεϊνης p16 σε τραχηλικά επιχρίσματα, ειδικά όταν αυτή είναι έντονη και εντοπίζεται τόσο στον πυρήνα όσο και στο κυτταρόπλασμα του κυττάρου, μπορεί να αποτελεί ένα χρήσιμο διαγνωστικό δείκτη για την τεκμηρίωση της παρουσίας υψηλού βαθμού (αντί για χαμηλού βαθμού) ενδοεπιθηλιακής αλλοίωσης, ιδιαιτέρως στις περιπτώσεις με αμφιλεγόμενα κυτταρομορφολογικά ευρήματα και γ) ότι η συνδυασμένη ανοσοκυτταροχημική χρώση για τις πρωτεϊνες p16 και L1 μπορεί επίσης να συμβάλλει στη διάκριση των χαμηλόβαθμων από τις υψηλόβαθμες αλλοιώσεις,και στην αναγνώριση των γυναικών υψηλού κινδύνου για την ανάπτυξη διηθητικής νόσου, ακόμη και σε περιπτώσεις με άγνωστο HPV status. Η πιθανή αξία των πρωτεϊνών που μελετήσαμε ως δεικτών πρόγνωσης των χαμηλού βαθμού ενδοεπιθηλιακών αλλοιώσεων δεν επιβεβαιώθηκε στο υλικό της παρούσας εργασίας
Bone marrow micrometastases in different solid tumors: Pathogenesis and importance
No full textEarly dissemination of cancer cells from the primary tumor via the circulatory system may result in the formation of microscopic metastatic deposits (micrometastases, MMs) in secondary compartments such as the bone marrow (BM), where there is a favorable environment for their subsequent growth and spread. MMs are considered the main reason for metastatic relapse in patients with early stage solid cancers after resection of the primary tumor. Although the molecular pathways leading to MMs remain only partly understood, there is increasing evidence that the detection of MMs in BM aspirates at the time of primary diagnosis is an independent prognostic factor, with a major influence in the stratification of these patients for adjuvant clinical treatment. Further potential applications of the detection of MMs include their use in monitoring therapeutic response or even in revealing targets for novel systemic therapies. All these intriguing possibilities are intensely investigated and carry great promise for radical improvements in the assessment and treatment of several epithelial. cancers which are currently to blame for the majority of cancer-related deaths in the industrialized world. (C) 2008 Elsevier Ltd. All rights reserved
Recent advances in the detection of bone marrow micrometastases: A promising area for research or just another false hope? A review of the literature
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