The Trojan horse Escherichia coli antibiotic microcin C (McC) consists of a heptapeptide attached to adenosine through a phosphoramidate linkage. McC is synthesized by the MccB enzyme, which terminally adenylates the ribosomally synthesized heptapeptide precursor MccA. The peptide part is responsible for McC uptake; it is degraded inside the cell to release a toxic nonhydrolyzable aspartyl-adenylate. Bionformatic analysis reveals that diverse bacterial genomes encoding mccB homologues also contain adjacent short open reading frames that may encode MccA-like adenylation substrates. Using chemically synthesized predicted peptide substrates and recombinant cognate MccB protein homologs, adenylated products were obtained in vitro for predicted MccA peptide-MccB enzyme pairs from Helicobacter pylori, Streptococcus thermophilus, Lactococcus johnsonii, Bartonella washoensis, Yersinia pseudotuberculosis, and Synechococcus sp. Some adenylated products were shown to inhibit the growth of E. coli by targeting aspartyl-tRNA synthetase, the target of McC.

We identified a novel human-specific family of transposable elements that consists of fused copies of the CpG-island containing the first exon of gene MAST2 and retrotransposon SVA. We propose a mechanism for the formation of this family termed CpG-SVA, comprising 5'-transduction by an SVA insert. After the divergence of human and chimpanzee ancestor lineages, retrotransposon SVA has inserted into the first intron of gene MAST2 in the sense orientation. Due to splicing of an aberrant RNA driven by MAST2 promoter, but terminally processed using SVA polyadenylation signal, the first exon of MAST2 has fused to a spliced 3'-terminal fragment of SVA retrotransposon. The above ancestor CpG-SVA element due to retrotranspositions of its own copies has formed a novel family represented in the human genome by 76 members. Recruitment of a MAST2 CpG island was most likely beneficial to the hybrid retrotransposons because it could significantly increase retrotransposition frequency. Also, we show that human L1 reverse transcriptase adds an extra cytosine residue to the 3' terminus of the nascent first strand of cDNA.

Dynactin is a multiprotein complex that enhances dynein activity. The largest dynactin subunit, p150Glued, interacts with microtubules through its N-terminal region that contains a globular cytoskeleton-associated protein (CAP)-Gly domain and basic microtubule-binding domain of unknown structure. The p150Glued gene has a complicated intron-exon structure, and many splice isoforms of p150Glued protein have been predicted. Here we describe novel natural 150 kDa isoforms: the p150Glued-1A isoform, whose basic domain is composed of 41 amino acids, and p150Glued-1B with a basic domain of 21 aa because of the lack of exons 5-7 in the corresponding messenger RNA (mRNA). According to reverse transcriptase-polymerase chain reaction (RT-PCR) and western blot data, p150Glued-1A is expressed in nerve tissues, in cultured cells and in embryonic tissues, while 1B is expressed ubiquitously. Overexpression of GFP-p150Glued-1A and -1B fusion proteins and immunostaining of cultured cells with 1A-specific antibodies show that the p150Glued-1A isoform is distributed along microtubules, whereas 1B is associated with microtubule plus-ends. The higher affinity of the p150Glued-1A isoform for microtubules is confirmed by a co-pelleting assay. In fibroblast-like cells, the interaction of p150Glued-1A with microtubules is less dependent on EB1/EB3 and CLIP170 proteins, compared with p150Glued-1B. In polarized cells, p150Glued-1A decorates microtubules that face the leading edge of the cell. The pattern of p150Glued-1A and p150Glued-1B interaction with microtubules and their tissue-specific expression patterns suggest that these isoforms might be involved in cell differentiation and proliferation.

Translation inhibitor microcin C (McC) is a heptapeptide with an aspartate α-carboxyl group linked to AMP via phosphoramidate bond. Modification of the McC phosphate by an aminopropyl moiety increases the biological activity by ~10-fold. Here, we determine the pathway of the aminopropylation reaction of McC. We show that the MccD enzyme uses S-adenosyl methionine to transfer 3-amino-3-carboxypropyl group onto a phosphate of an McC maturation intermediate consisting of adenylated heptapeptide. The carboxyl group is removed by the MccE enzyme, yielding mature McC. MccD is an inefficient enzyme that requires for its action the product of Escherichia coli mtn gene, a 5'-methylthioadenosine/S-adenosylhomocysteine nucleosidase, which hydrolyses 5'-methylthioadenosine, the product of MccD-catalyzed reaction, thus stimulating the amino-3-carboxypropylation reaction. Both MccD and MccE are capable of modifying McC-like compounds with divergent peptide moieties, opening way for preparation of more potent peptidyl-adenylates.