Translational
Frameshift may be Occur in p11, an Interaction Protein of Cx31, in Yeast
HUANG Liang-Qun, SHI Xiao-Liu, TAN Zhi-Ping,
FANG Hai-Yan, ZHENG Duo, XIA Jia-Hui, XIA Kun*
( National Laboratory of Medical Genetics of China, Changsha 410078,
China )
Abstract Connexin 31
is a member of connexins family. The carboxy-terminal cytosolic domain of
connexin 31 contains several potential phosphorylation sites. In this work, a
yeast two-hybrid protein interaction screen have been used to identify proteins
that bind to the carboxy-terminus of connexin 31, and the p11 protein, an
unique member of S100 protein family, and one of the two subunits of the
annexin II tetramer was isolated. Interestingly, from yeast two-hybrid AD’s
coding sequence, three different reading frames of p11 DNA sequence were
found,which come from different AD plasmids. By constructing AD plasmids using
p11 ORF or 5＇ UTR, the protein coding by p11 ORF bind
to connexin 31, while polypeptides coding by three kinds of 5＇UTR did not bind to connexin
31, suggesting atranslational frameshift of p11 fusion protein. To construct
baits by deviding connexin 31 C-terminus into two domain, the p11 binding
domain of connexin 31 was found located between 206―237 codons. The plasmid
Cx31CT-pGEX-4T-2 was constructed for expression and purification of GST-Cx31CT;
and p11-pQE30 for expression and purification of 6×His-p11. In vitro
binding assay showed that recombinant Cx31 interacted with recombinant p11.
Key words connexin 31; yeast two-hybrid; calpaction I;
light chain; frameshift
*Corresponding author: Tel,
86-0731-4472093; e-mail, [email protected]