Table 1.
List of primers used in this study.
Table 2.
List of strains and plasmids used in this study.
Figure 1.
Overview of the turnerbactin biosynthetic gene cluster.
A. Organization of genes involved in turnerbactin biosynthesis. The locations of putative Fur boxes are indicated by asterisks (*). The black triangle indicates the location of NRPS disruption. B. Domain organization of the NRPS modules of TnbF. C, condensation domain; A, adenylation domain; PCP, peptidyl-carrier protein; TE, thioesterase. C. Putative Fur box sequences of turnerbactin's biosynthetic gene cluster compared to the bacillibactin biosynthetic operon of Bacillus subtilis. Shaded bases match the revised view of Fur box sequences consisting of two overlapping 7–1–7 motifs proposed by Baichoo and Helmann.
Table 3.
Proteins with similarity to the products of the turnerbactin biosynthetic gene cluster. S: similarity, I: identity.
Figure 2.
Fe(III)-binding activity of T. turnerae T7901 cultures.
The CAS assay was used to measure Fe(III)-binding activity of iron-replete and iron-limited culture supernatants. Iron-replete conditions contained 10 μM ferric iron, while iron-limited conditions contained 0.1 μM ferric iron.
Figure 3.
Structures of siderophores isolated from T. turnerae T7901.
Table 4.
Molecular ions and common mass fragments of siderophore from T. turnerae T7901. Fragment losses refer to the compound listed immediately above in the table.
Table 5.
NMR data for 1 and 3 (800 MHz) in CD3OD.
Figure 4.
Fe(III)-binding activity of wild-type T. turnerae T7901 compared to tnbF mutant TtAH03.
Disruption of tnbF leads to a significant decrease in siderophore activity, as measured by the CAS assay. CAS activity was normalized to OD600 measurements of each culture.
Figure 5.
Comparison of wild-type T7901 and TtAH03 extracts by HPLC, recorded at 215 nm.
Figure 6.
Proposed biosynthesis of turnerbactin.
Bold structures indicate the most recent addition to the compound.
Figure 7.
HPLC, HRMS, and MS/MS analysis of L. pedicellatus extracts.
A. Extract of L. pedicellatus. B. Standard of DHB-Orn-Ser, isolated from T. teredinibacter T7901. Inset in both figures shows the HRMS of the peak indicated. The shipworm extract shows other compounds with similar mass, but also contains a compound with nearly identical high resolution mass as the DHB-Orn-Ser standard of 356.1446. Red boxes indicate shared MS/MS fragmentation peaks of the 356 molecular ion between the shipworm extract and the siderophore standard.