Part:BBa_K5398601

Mfp6

Mussels foot protein type 6 (Mfp6), with a molecular weight of 13.8 kDa, is a protein found in the byssal gland cells of mussels. It belongs to the family of mussel foot proteins and plays a crucial role in maintaining the reducing conditions necessary for optimal wet adhesion in marine Mussels. Mfp6 is particularly rich in cysteine, a sulfur-containing amino acid that can form stable structures in mussel adhesion proteins and provide antioxidant protection. This antioxidant property helps to prevent the over-oxidation of dopamine (DOPA) residues in adhesion proteins, thus maintaining their adhesive function.

During the adhesion process of mussels, Mfp6 may work in conjunction with other foot proteins, such as Mfp3 and Mfp5, which are rich in dopamine and are key factors in the adhesive strength of mussel adhesion proteins. Mfp6, through its antioxidant properties, helps to protect dopamine residues at the initial stage of adhesion, thereby maintaining the protein's adhesive ability. In addition, Mfp6 may also be involved in regulating the tautomeric balance of adhesion proteins, further affecting adhesion performance.

To validate the functionality of Mfp6, we designed bacteria expressing Mfp6.We constructed the pET28a (+) vector, after culturing at 37°C for 5 hours, extracted the proteins for SDS-PAGE and Coomassie Brilliant Blue staining analysis.

Fig. 1 Protein pre-expression of SUMO-Mfp6(28 kDa). Lane 1: Marker. Lane 2: Mfp6-Whole Cell Lysate(+IPTG). Lane 3: Mfp6-Supernatant(+IPTG). Lane 4: Mfp6-Pellet-PBS(+IPTG). Lane 5: Mfp6-Pellet-Extraction buffer(+IPTG). Lane 6: Mfp6-Whole Cell Lysate-1(CK). Lane 7: Mfp6-Supernatant-1(CK). Lane 8: Mfp6-Pellet-PBS-1(CK). Lane 9: Mfp6-Pellet-Extraction buffer-1(CK). Lane 10: Mfp6-Whole Cell Lysate-2(CK). Lane 11: Mfp6-Supernatant-2(CK). Lane 12: Mfp6-Pellet-PBS-2(CK). Lane 13: Mfp6-Pellet-Extraction buffer-2(CK).

We purified SUMO-Mfp6 using a HiTrap Ni-NTA column. The purified protein was verified by SDS-PAGE and was found to be present in the XX mM imidazole elution fraction.

Fig. 2 Protein expression of SUMO-Mfp6(28 kDa).

We used a microplate reader to assay the activity of Mfp6. Fig. 2 illustrates the functionality of Mfp6. The dopamine content without Mfp6 was XX, and the dopamine content with the addition of Mfp6 was XX. These results indicate that Mfp6 significantly increases the retention of dopamine, suggesting its role in stabilizing dopamine against oxidation.

Fig. 3 Assay of Mfp6 activity.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]