A novel expression system for recombinant marine mussel adhesive protein Mefp1 using a truncated OmpA signal peptide.
To express an increased level of recombinant Mefp1 (marine mussel adhesive protein) in soluble form, we constructed expression vectors encoding truncated OmpA signal peptide-Mefp1 fusion proteins. OmpA signal peptide (OmpASP) is the 21 residue peptide fragment of the 23 residue OmpA signal sequence cleavable by signal peptidase I. We successfully produced increased levels of soluble recombinant Mefp1 (rMefp1) with various deletions of OmpASP, and found that the increased expression was caused by the increased pI of the N-terminus of the fusion proteins (> or = 10.55). All the OmpA signal peptide segments of 3-21 amino acids in length had the same pI value (10.55). Our results suggest that the pI value of the truncated OmpASP (OmpASP(tr)) play an important role in directional signaling for the fusion protein, but we found no evidence for the presence of a secretion enhancer in OmpASP. For practical applications, we increased the expression of soluble rMefp1 with OmpASP(tr) peptides as directional signals, and obtained rMefp1 with the native amino terminus (nN-rMefp1) using an OmpASP(tr)| Xa leader sequence that contains the recognition site for Xa protease.
Lee
Sang Jun
SJ
Biotechnology Research Institute, National Fisheries Research and Development Institute, Busan 619-902, Korea. sjlee@nfrdi.re.kr
Han
Yun Hee
YH
Nam
Bo Hye
BH
Kim
Young Ok
YO
Reeves
Peter
P
D000595:Amino Acid Sequence; D000818:Animals; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics*; D001483:Base Sequence; D015153:Blotting, Western; D004926:Escherichia coli / Q000235:genetics*; D015870:Gene Expression; D005822:Genetic Vectors; D008969:Molecular Sequence Data; D021382:Protein Sorting Signals / Q000235:genetics*; D011506:Proteins / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics*; D014443:Tyrosine / Q000378:metabolism
D001425:Bacterial Outer Membrane Proteins; D021382:Protein Sorting Signals; D011506:Proteins; D011993:Recombinant Fusion Proteins; C037403:adhesive protein, mussel; C078167:OMPA outer membrane proteins; D014443:Tyrosine