INVESTIGADORES
LONGHI Silvia Andrea
artículos
Título:
Interaction map of the Trypanosoma cruzi ribosomal P protein complex (stalk) and the Elongation Factor 2
Autor/es:
SMULSKI CR*; LONGHI SA*; JURI AYUB M; EDREIRA MM; SIMONETTI L; GOMEZ KA; BASILE JN; CHALOIN O; HOEBEKE J; LEVIN MJ
Revista:
JOURNAL OF MOLECULAR RECOGNITION
Editorial:
JOHN WILEY & SONS LTD
Referencias:
Lugar: Washington; Año: 2011 p. 359 - 370
ISSN:
0952-3499
Resumen:
The large subunit of the eukaryotic ribosome possesses a long and protruding stalk formed by the ribosomal P
proteins. This structure is involved in the translation step of protein synthesis through interaction with the elongation
factor 2 (EF-2). The Trypanosoma cruzi stalk complex is composed of four proteins of about 11 kDa, TcP1a, TcP1b,
TcP2a, TcP2b and a fifth TcP0 of about 34 kDa. In a previous work, a yeast two-hybrid (Y2H) proteinprotein
interaction map of T. cruzi ribosomal P proteins was generated. In order to gain new insight into the assembly of the
stalk, a complete interaction map was generated by surface plasmon resonance (SPR) and the kinetics of each
interaction was calculated. All previously detected interactions were confirmed and new interacting pairs were found,
such as TcP1bTcP2a and TcP1bTcP2b. Moreover P2 but not P1 proteins were able to homo-oligomerize. In addition,
the region comprising amino acids 210270 on TcP0 was identified as the region interacting with P1/P2 proteins,
using Y2H and SPR. The interaction domains on TcP2b were also mapped by SPR identifying two distinct regions. The
assembly order of the pentameric complex was assessed by SPR showing the existence of a hierarchy in the
association of the different P proteins forming the stalk. Finally, the TcEF-2 gene was identified, cloned, expressed and
refolded. Using SPR analysis we showed that TcEF-2 bound with similar affinity to the four P1/P2 ribosomal P proteins
of T. cruzi but with reduced affinity to TcP0.