## 3 Results and Discussion
### Analysis of Plasmid Presence and Antibiotic Resistance Genes
#### 3.6.3 Lanthopeptide and RiPP Gene Clusters
In summary, BGC region 21 contains a cryptic Liy gene cluster involved in the biosynthesis of class V lanthippeptides. This cluster includes regulatory genes (XJJ18547, LivR1; XJJ18548, LiyR2) and transport-related genes such as XJJ18563 (LiyT3), highlighting the region's potential for producing novel antibiotics (Figure 5, Table S17). BGC Region 22 Derived from _Lentza_ sp. JNUCC 0626  
The antiSMASH analysis of BGC region 22 identified the XJJ18652 protein as a core biosynthetic gene, implicating its involvement in a RiPP biosynthetic cluster (Figure S10, Table S18). Additionally, the XJ18652 protein was found to encode a member of the domain of unknown function 692 (DUF692), an emerging family of enzymes associated with post-translational modifications during the biosynthesis of RiPPs. These enzymes are characterized by multinuclear iron centers, with only two family members, MbnB and TglH, functionally characterized to date. A recent study has proposed reclassifying this family as multinuclear non-heme iron-dependent oxidative enzymes (MNIOs) [50]. As expected, structural predictions using HHPred and SWISS-MODEL indicated that the XJ18652 protein shares the highest similarity with the methanobactin biosynthetic protein MbnB from Methylosinus trichosporum (PDB: 7 TCX), MbnB from _Vibrio caribbeanicus_ (PDB: 7 DZ9), and the RiPP recognition protein TglH involved in peptidyl (S)-2-mercaptoethanolglycine biosynthesis from _Pseudomonas syringae_ (PDB: 8H7). The intriguing aspect of BGC region 22 is the presence of four consecutive TIGR04222 domain-containing membrane proteins located in an operon-like arrangement within the DUF692 downstream. Notably, the proteins XJ1J8648, XJ1J8649, XJ1J8650, and XJ1J8651 each exhibit Cys- and Ser-rich C-terminal sequences, as follows: \"C  
protein from Neisseria gonorrhoeae (PDB: 3 DEE). In contrast, protein XJIJ18654 encodes an S8 family serine peptidase. S8 family serine peptidases are critical in the lantipeptide biosynthetic pathway, where they facilitate the precise processing and maturation of precursor peptides, a process essential for the production of biologically active lantiopeptides [52; 53]. BGC Region 25 Derived from _Lentzea_ sp. JNUCC 0626  
In BGC region 23, a class III lanthionine biosynthetic gene cluster (BGC) comprising six proteins was identified using the antiSMASH approach (Figure S11, Table S19). Proteins XJIJ19011 (852 aa) and XJIJ19012 (55 aa) exhibit significant sequence similarity to the class III lanthionine synthetase LanKC and the SapB/AmfS family lanthipeptides, as determined by BLASTP analysis. Additionally, proteins XJIJ19013 (538 aa) and XJIJ19014 (572 aa), which are ABC transporter ATP-binding proteins, are positioned in the same transcriptional direction in their upstream regions. Conversely, protein XJIJ19015 (167 aa), which shares high similarity with DUF3145 domain-containing proteins, is oriented in the opposite transcriptional direction. However, protein XJIJ19016 (440 aa), exhibiting similarity to S8 family serine peptidases, is located in the upstream region. Based on these observations, the minimal unit of the class III lanthionine BGC in region 25 is inferred to consist of six proteins. Notably, in class III BGCs, S8 family serine peptidases play a critical role in the processing of precursor peptides. These peptidases cleave the precursor peptides at specific sites to release mature lanthipeptides, a process that is essential for the proper maturation and biological activity of the lanthipeptides.