A type IC restriction-modification system in Lactococcus lactis

Abstract

Three genes coding for the endonuclease, methylase, and specificity subunits of a type I restriction-modification (R-M) system in the Lactococcus lactis plasmid pIL2614 have been characterized. Plasmid location, sequence homologies, and inactivation studies indicated that this R-M system is most probably of type IC.

FIG. 1

(Top) Organization of the pIL2614 sequenced fragment. A putative promoter sequence is indicated by an arrow, and a transcription terminator is indicated by a circle atop a vertical line. (Bottom) Features of the ORFs. Start and stop numbers refer to positions in the sequence, the corresponding codons are shown in parentheses. The putative ribosome binding site and the beginning of the ORFs are shown in capital letters.

FIG. 2

Alignment of the predicted amino acids of the pIL2614 HsdR peptide with the R subunits of EcoR124II (8), M. pulmonis (7), and H. influenzae (10). Sequence accession numbers are U90222, X13145, L25415, and L45919, respectively. Helicase-like domains I to VI (14) and X and Y domains (40) as well as the additional conserved domain Z are shown in boldface letters. Conserved amino acids and conservative or semiconservative substitutions are indicated by an asterisk and a period, respectively.

FIG. 3

Alignment of the predicted amino acids of the pIL2614 HsdM peptide with the M subunits of EcoR124II (21) and M. pulmonis (7) (GenBank accession no. U90222, X13145, and L25415, respectively). Conserved amino acids and conservative or semiconservative substitutions are indicated by an asterisk and a period, respectively. Conserved motifs (CMIs, CMI, CMII, and CMIII) are shown in boldface letters, and the proposed consensus for the N12 class and all MTases (39) are indicated below. Different groups of amino acids are indicated as follows: p, polar (D, E, N, H, K, R, S, Q, and G); h, hydrophobic (W, F, I, L, M, V, A, P, Y, C, and T); n, negatively charged (D and E); f, aromatic (F, W, Y, and H); a, aliphatic (I, L, V, and M); c, charged (D, E, K, R, and H); and s, small, nonbulky (G, A, S, T, D, N, P, and V).

FIG. 4

Predicted amino acid sequence of the pIL2614 HsdS peptide. Thirty-nine-amino-acid repeats (A and A′) and split repeats (D and D′) (25) are shown in boldface letters. The consensus for the 24-aa repeats present in all S peptides (21) is indicated below, together with sequences from EcoR124II (21), M. pulmonis (7), and S. citri (27).