Nucleic Acids Research Overproduction and purification of the connector protein of Bacillus subtiis phage 029 Carmen IbaMiez, Juan A.Garcia, Jose L.Carrascosa and Margarita Salas Centro de Biologia Molecular (CSIC-UAM), Universidad Aut6noma, Canto Blanco, Madrid-34, Spain Received 19 December 1983; Revised and Accepted 17 February 1984 ABSTRACT A 029 DNA fragment containing genes 10 and 11, coding for the connector protein and the lower collar protein, respectively, has been cloned in the pBR322 derivative plasmid pKC30 under the control of the PL promoter of phage lambda. Two polypeptides with the electrophoretic mobility of proteins plO and pll were label- led with 35S-methionine after heat induction. The proteins were characterized as plO and pll by radioimmunoassay and they repre- sented about 10% and 7%, respectively, of the total E. coli pro- tein after 4 hours of induction. These proteins represent less than 1% of the B. subtilis protein in 029-infected cells. Protein plO has been highly purified from the E.coli cells carrying the recombinant plasmid. Antibodies raised against the purified pro- tein plO reacted with the connector protein produced in 029-in- fected B. subtilis. INTRODUCTION The head-tail connecting region in bacteriophages contains a protein, the connector, that plays a key role in the early steps of prohead assembly. It has been shown recently in the B. subtilis phage 029 (1,2) and in the E. coli phages T4 (3) and lambda (4) that the connector is an oligomer of a single protein with 12-fold symmetry. This is, therefore, the region at which the symmetry transition between the lower corner of the phage head (5-fold symmetry) and the tail (6-fold symmetry) occurs. According to Hendrix (5), the connector is also an active device moving the DNA into the phage prohead. In addition, in phage 029, the connector is essential for the normal prohead elongation (6,7). Neck-tail complexes of phage 029, composed of proteins plO (upper collar or connector), p1l (lower collar), p12* (neck ap- pendages) and p9 (tail), have been isolated and the symmetry of the neck region (proteins plO and pll) determined (1,2). To stu- dy the structure of the isolated connector protein, plO, and to Volume 12 Number 5 1984 0 1 R L Press Limited, Oxford, England. 2351