TY - JOUR
T1 - Polynucleotide phosphorylase interacts with ribonuclease E through a ββαββα domain
AU - Durán-Figueroa, Noé V.
AU - Piña-Escobedo, Alberto
AU - Schroeder, Imke
AU - Simons, Robert W.
AU - García-Mena, Jaime
N1 - Funding Information:
This work was partially supported by UC Mexus-CONACYT and CONACYT (36465-N) grants. We thank Sidney R. Kushner for pQLK16 plasmid, to Agamemnon J. Carpousis for anti-RNase E, and to Kaj Frank Jensen for anti-RNase PH antibodies. We thank Cecília M. Arraiano, Monica Amblar Esteban, Evgeny A. Nudler, and Paul B. Fisher for discussion and suggestions on this work. We also thank Ben F. Luisi, and Gianni Dehò for critical reading of this manuscript; Martyn F. Symmons for support on protein structure analysis; to Carolina Miranda-Brito for PCR-cloning; to Guadalupe Aguilar-Gonzalez for DNA sequencing; to Rodrigo García-Gutiérrez for clerical and to Antonia López-Salazar for administrative assistance. During this work NVDF was partially supported by fellowships from CONACYT (Grant 36465-N) and PRONABES-IPN.
PY - 2006/6
Y1 - 2006/6
N2 - In the present work we have used a double-hybrid assay in bacteria to identify a putative domain in E. coli PNPase required for in vivo interaction with RNase E. We used a 202 aa fragment of RNase E previously reported as the PNPase binding domain in this enzyme and a collection of 13 different fragments of 105 aa, spanning the entire sequence of 734 aa PNPase (GenBank Accession number NP_417633). Our results indicate that two clones of PNPase including residues 158-262 and residues 473-577 contain interaction sites for RNase E within a ββαββα domain configuration. Three-dimensional modeling of the E. coli PNPase based on the S. antibioticus protein structure indicates that the putative binding domain is located on the monomer surface, facing outward from the trimeric tertiary structure. Since a copy of the ββαββα domain is also found in RNase PH, we investigated and found an interaction with RNase E in a pull-down assay. We suggest this interaction takes place through the similar ββαββα domain present in the tertiary structure of this enzyme. Based on these results, we propose that RNase PH and RNase E could form functional assemblies in E. coli.
AB - In the present work we have used a double-hybrid assay in bacteria to identify a putative domain in E. coli PNPase required for in vivo interaction with RNase E. We used a 202 aa fragment of RNase E previously reported as the PNPase binding domain in this enzyme and a collection of 13 different fragments of 105 aa, spanning the entire sequence of 734 aa PNPase (GenBank Accession number NP_417633). Our results indicate that two clones of PNPase including residues 158-262 and residues 473-577 contain interaction sites for RNase E within a ββαββα domain configuration. Three-dimensional modeling of the E. coli PNPase based on the S. antibioticus protein structure indicates that the putative binding domain is located on the monomer surface, facing outward from the trimeric tertiary structure. Since a copy of the ββαββα domain is also found in RNase PH, we investigated and found an interaction with RNase E in a pull-down assay. We suggest this interaction takes place through the similar ββαββα domain present in the tertiary structure of this enzyme. Based on these results, we propose that RNase PH and RNase E could form functional assemblies in E. coli.
KW - Polynucleotide phosphorylase
KW - Protein interaction
KW - Ribonuclease E
KW - Ribonuclease PH
KW - ββαββα domain
UR - http://www.scopus.com/inward/record.url?scp=33745955546&partnerID=8YFLogxK
U2 - 10.1016/j.biochi.2006.01.002
DO - 10.1016/j.biochi.2006.01.002
M3 - Artículo
SN - 0300-9084
VL - 88
SP - 725
EP - 735
JO - Biochimie
JF - Biochimie
IS - 6
ER -