Linaridins and lanthipeptides are two classes of natural products belonging to the ribosomally synthesized and posttranslationally modified peptide (RiPP) superfamily. Although these two RiPP classes share similar structural motifs such as dehydroamino acids and thioether-based cross-links, the biosynthesis of linaridins and lanthipeptides involved distinct sets of enzymes. Here, we report the identification of a novel lanthipeptide cypepeptin from a recombinant strain of Streptomyces lividans, which harbors most of the cypemycin (a prototypic linaridin) biosynthetic gene cluster but lacks the decarboxylase gene cypD. In contrast to the generally believed structure of cypemycin, multiple d-amino acids and Z-dehydrobutyrines were observed in both cypepeptin and cypemycin, and the stereochemistry of each amino acid was established by the extensive structural analysis in combination with genetic knockout and mutagenesis studies. Comparative analysis of cypemycin and cypepeptin showed that the aminovinyl-cysteine (AviCys) moiety of cypemycin plays an essential role in disrupting the cell integrity of M. luteus, which cannot be functionally substituted by the structurally similar lanthionine moiety.