Factors limiting display of foreign peptides on the major coat protein of filamentous bacteriophage capsids and a potential role for leader peptidase

Abstract

Many small peptides can be displayed on every copy of the major coat protein in recombinant filamentous bacteriophages but larger peptides can only be accommodated in hybrid virions mixed with wild-type protein subunits. A peptide insert of 12 residues capable of display at high copy number in a hybrid virion was found to be incapable of supporting recombinant virion assembly, a defect that could not be overcome by over-expressing leader peptidase in the same Escherichia coli cell. In contrast, over-expressing leader peptidase did increase the copy number of two 9-residue peptides that were poorly incorporated into hybrid virions. The factors that limit peptide display are varied and not restricted to the early stages of viral assembly.z 1998 Federation of European Biochemical Societies.

Phage display particles expressing tumor‐specific antigens induce preventive and therapeutic anti‐tumor immunity in murine p815 model

“…The pVIII of filamentous phage is synthesized with a leader sequence required to insert pVIII across the membrane; the leader sequence is then cleaved off by the bacterial leader peptidase. 35 Although 6 ϫ Histidine tag is usually placed at the C-terminus to prevent it from interfering with the signal sequence, our data showed the location of 6 ϫ Histidine tag between the leader sequence and mature pVIII did not have an effect on assembly of phage virion. The direct ELISA assay using anti-His6 MAb demonstrated the consecutive Histidine tagged phage particles can be purified on Ni-NTA agarose column (Fig.…”

“…To determine whether the P1A display phage particle was able to induce a CTL response against antigen, P815A in vivo, P1A [35][36][37][38][39][40][41][42][43] specific CTL responses in immunized mice were determined by release of Cr 51 We observed P1A [35][36][37][38][39][40][41][42][43] -specific CTL responses in almost all the mice that injected the recombinant phages (pC89hisP1A). The results of the experiment are shown in Figure 3, the splenocyte cultures being derived from mice immunized with pC89hisP1A exhibited lytic activity against P815 (Fig.…”

“…FIGURE 4 -The recombinant phage particles administered induce a Th1-like response. DBA/2 mice were primed s.c. into the base of tail with phage display particles pC89hisP1A, control phage particles pC89hisOVA or peptide P1A [35][36][37][38][39][40][41][42][43] . Fourteen days later, spleen cells were stimulated with phage display particles pC89hisP1A and peritoneal macrophage in vitro and then the supernatant were tested for cytokines IFN-␥ and IL-4 (solid bars).…”

“…These data showed that injection of hybrid phage particles into DAB/2 mice prevented or suppressed tumor growth. Additionally, immunization primed a P1A [35][36][37][38][39][40][41][42][43] -specific CTL response and promoted phage particle-associated Th1 cytokine response. The potential of developing anti-tumor vaccine with filamentous phage particles is discussed.…”

Generation of Metal-Binding Staphylococci through Surface Display of Combinatorially Engineered Cellulose-Binding Domains

“…Gram-positive surface display systems have been suggested to exhibit some advantages compared to gram-negative bacteria (28,49): (i) the translocation involves only a single membrane, and (ii) gram-positive bacteria have been shown to be more rigid and thus less sensitive to shear forces (19,36) due to the thick cell wall surrounding the cells, and thus potentially more suitable for field applications such as bioadsorption. For metal adsorption applications, gram-positive bacteria have the additional advantage of having inherent metal-binding capacity due to the thick peptidoglycan layer (31).…”