3626 TDFLRMMLQEER(8/16) 1 Phage display (common panning) 3 PMID:35534543 Anti-interferon gamma (IFN-gamma) monoclonal antibody B27 Interferon gamma, IFN-gamma Not determined. Not determined. SUT12 M13 phage display library ELISA Their binding activity against B27 mAb was detected by phage ELISA. Data were not shown. The specific peptide sequence recognized by B27 mAb, TDFLRMMLQEER, was retrieved from a phage display random peptide library. Sequence alignment and homology modeling demonstrated that the queried phage peptide sequence and structure were similar to amino acids at position 27–40 (TLFLGILKNWKEES) of the human IFN-γ. 3627 DYHDPSLPTLRK(4)[0.7929±0.1464, NT] GNNPLHVHHDKR(2)[0.6135±0.0746, NT] STVRPLLMMDKY(1)[1.2004±0.2109, NT] HHLRIPYALDQT(1)[0.5489±0.0904, NT] DSAPSYNYRPSY(1)[1.6395±0.1421, 520±7] DYHDPSPPTLRK(1)[0.6436±0.2095, NT] KVYFSIPWRVPM(1)[1.6281±0.198, 74±3] QVNGLGERSQQM(1)[1.1559±0.1119, NT] HSNDPRLITMRK(1)[0.3179±0.2239, n.d.] TCFAHTHNNFGH(1)[0.8001±0.3975, NT] RDYHPRDHTATW(1)[0.3394±0.1579, NT] IPGTAPPLARTG(1)[1.044±0.023, NT] KDFLPSPQTATW(1)[1.5606±0.1277, 488±33] VRAFSGEHSFVS(1)[1.1746±0.023, NT] 14 Phage display (common panning) 3 PMID:35059169 9-fluorenylmethoxycarbonyl-phenylalanine-tyrosinephosphate (Fmoc-Fργ ) micellar aggregates Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) ELISA The absorbance at 405 nm was monitored in a SpectraMax i3 (Molecular Devices). Wild-type M13 phage has been used as a positive control of ELISA assay and as a negative control of Fmoc-FpY binding. In addition, NMR experiments were acquired in a Bruker Avance III 600 spectrometer equipped with a triple-resonance cryoprobe (TCI). Spectra were processed using software TOPSPIN (Bruker Biospin, Karlsruhe, Germany) and peaks assigned with Sparky (TD Goddard and DG Kneller, Sparky 3, University of California, San Francisco, USA). The dissociation constants KD (μM) were determined in the OriginPro9. The first column of affinity values is the absorbance405 value reproduced from Figure 1C, the second column is the KD (μM) determined by NMR experiments. The lead peptide, KVYFSIPWRVPM-NH2 (P7) was found to bind to the Fmoc-FpY ligand exclusively in its self-assembled state with KD =74 ± 3 μM. Circular dichroism, NMR and molecular dynamics simulations revealed that the peptide interacts with Fmoc-FpY through the KVYF terminus and this binding event disrupts the assembled structure. In absence of the target micellar aggregate, P7 was further found to dynamically alternate between multiple conformations, with a preferred hairpin-like conformation that was shown to contribute to supramolecular ligand binding. Three identified phages presented appreciable binding, and two showed to catalyze the hydrolysis of a model para-nitro phenol phosphate substrate, with P7 demonstrating conformation-dependent activity with a modest kcat/KM = 4 ± 0.3 * 10^(-4) M^(-1)s^(-1). 3628 ACVWWDHTYC(8/19)[2.0156±0.0932] ACNWWDLTLC(6/19)[1.8097±0.0919] ACTWWDMAFC(4/19)[1.8452±0.1287] ACEWWDVTYC(1/19)[2.5513±0.0711] 4 Phage display (subtractive panning) 3 PMID:37126866 Anti-CPA monoclonal antibody MAB-1418 Cyclopiazonic acid, CPA Not determined. Not determined. Ph.D.-C7C phage display library (CX7C) ELISA The analytical signal was obtained by measuring the absorbance at 450 nm with a CLARIOstar microplate reader from BMG (Ortenberg, Germany). Data shown were reproduced from Figure 2. A pre-selection step against BSA was carried out to eliminate non-specific phages prior to the actual selection with mAb-1418 in each round. The mimotope sequence, ACNWWDLTLC, performed best in phage-based immunoassays, surface plasmon resonance binding analyses, and UC-RET-based immunoassays. To develop a homogeneous assay, upconversion nanoparticles (UCNP, type NaYF4:Yb3+, Er3+) were used as energy donors and coated with streptavidin to anchor the synthetic biotinylated mimotope. Alexa Fluor 555, used as an energy acceptor, was conjugated to the anti-CPA antibody fragment. The homogeneous single-step immunoassay could detect cyclopiazonic acid (CPA) in just 5 min and enabled a limit of detection (LOD) of 30 pg/mL (1.5 μg/kg) and an IC50 value of 0.36 ng/mL. No significant cross-reactivity was observed with other co-produced mycotoxins. Finally, we applied the novel method for the detection of CPA in spiked maize samples using high-performance liquid chromatography coupled to a diode array detector (HPLC-DAD) as a reference method. 3629 AGKCCFATGGS(5) ARKCCYGRGGS(4) ASRKCCYVGGS(2) AGGKCCYSGGS(1) AGKCCYSKGGS(1) AGKCCYSTGGS(1) AKCCYAGRGGS(1) AKCCYAHIGGS(1) ANRKCCFAGGS(1) ARAKCCYYGGS(1) ASKCCYHVGGS(1) AYKCCYWTGGS(1) ASSFLEQDGGS(1) AILIPFWTGGS(1) AIPYCMGVGGS(1) AWLRSMKVGGS(1) ANTGWQSRGGS(1) AVFFGVGWGGS(1) AKKVWIIMGGS(1) AMMTSSGGGGS(1) AYFVVRGLGGS(1) KCC[FY] 21 Phage display (common panning) 3-4 PMID:35665749 Zc RNase P ribozyme (RPR) Not determined. Not determined. Not determined. AXXXXXXXGGS phage display library We report that a hydrophobic-cationic RNA binding peptide selected by phage display (P43: AKKVWIIMGGS) forms insoluble amyloid-containing aggregates, which reversibly accrete RNA on their surfaces in an RNA-length and Mg2+- concentration dependent manner. The aggregates formed by P43 or its sequence-simplified version (K2V6: KKVVVVVV) inhibited RNA polymerase ribozyme (RPR) activity at 25mM MgCl2, while enhancing it significantly at 400mM MgCl2. Our work shows that such hydrophobic-cationic peptide aggregates can reversibly concentrate RNA and enhance the RPR activity, and suggests that they could have aided the emergence and evolution of longer and functional RNAs in the fluctuating environments of the prebiotic earth. 3630 RRPPR(5/24) 1 Phage display (common panning) 6 PMID:34111267 Rat heart microvascular endothelial cell, RHMVEC Not determined. Not determined. Not determined. Novagen T7 X7 phage display library Rat heart microvascular endothelial cell (RHMVEC) (80% confluent; approximately 2.0e7 cells / 100 mm dish) were washed with PBS and preincubated in serum-free medium at 37°C for 30 minutes and inoculated with an extract (5.0e9 pfu) of the T7 phage library to reach a multiplicity of infection (MOI) of 250. After incubation for 1 hour at 37°C, cells were washed with ice-cold PBS and acid washed with 0.1N HCl pH 2.2 for 15 seconds to remove unbounded and weakly associated phages from the cell surface. Cells were then trypsinized, centrifuged, and lysed with sterile deionized water on ice and internalized phages were amplified. After completion of 6 rounds of selection/amplification, Escherichia Coli BL21 was infected with the resulting phages and plated, individual plaques were picked, amplified, and sequenced. Phage display technology was used to identify a small peptide (RRPPR) that was internalized into endothelial cells. This peptide was potent in blocking nitric oxide (NO) release. Fusing RRPPR with a minimal Cav inhibitory domain (CVX51401) dose-dependently blocked NO release, VEGF induced permeability, and retinal damage in a model of uveitis. 3631 AYPGLPRSQS[155] AYPGPRTGHW[inactive] AYPGPHTFRG[inactive] AYPGWTTKVA[53] AYPGWPLRNA[61] AYPGWLVKNG[4.8] 6 Phage display (competitive panning) 3 PMID:35343875 Membranes of HEK293 cells expressing protease-activated receptor 4 (PAR4), HEK293-PAR4 Not determined. Not determined. Not determined. AYPGX6 phage display library Changes in fluorescence intensity were measured using FLIPR Tetra Systems. EC50 (μM) from the individual fits were calculated and shown. To obtain binding peptides by phage display, we opted to pan with membranes derived from HEK293 cells transiently expressing PAR4 receptor (HEK293-PAR4). The first panning cycle was carried out by incubating an approximately 10^10 library phage in a well coated with HEK293-PAR4 membranes. After 5 rinses and 1 x 5-minute wash with PBST (phosphate buffer saline containing 0.1% Tween 20), binding phage were then recovered and amplified by infection with log phage E. coli strain ER2738 (New England Biolabs). The second round of panning was carried out similarly to the first cycle, except that: (1) steps were carried out in duplicate wells so that binding phage could be recovered from the first well by infection and from the second well by elution with 200 μM AYPGKF peptide; (2) the panning step was preceded by 3 successive 30-minute pre-adsorptions in wells coated with wild-type HEK293 membranes; (3) amplified output phages from the first round were mixed with 0.2 μg of αVβ3 integrin (Chemicon, Burlington, MA) so that a competitive depletion of integrin-binding phage was possible in the panning step; and (4) wash steps included 5 fast rinses, 2 x 5-minute washes, and a further fast rinse. The third panning cycle was as cycle 2 except that: (1) 5 x 15-minute pre-adsorptions with HEK293 membranes were carried out; and (2) elution with ER2738 was replaced by a 10-minute elution with 0.1 M glycine buffer, pH 2.2, followed by neutralization with 0.1 vol of 2M Tris, pH 8.0. Throughout the steps, membranes were immobilized for 2 hours in wells of 96-well plates coated with wheat-germ agglutinin (Perkin Elmer, Waltham, MA) and blocked for 1.5 hours with PBS plus milk (5% fat free), followed by 4 rinses with PBS. All phage incubations were performed for 2 x 2 hours in PBS plus milk. All steps were carried out at 4℃ in the presence of a protease inhibitor cocktail. Using an AYPG-based biased phage-display peptide library approach followed by chemical peptide optimization, A-Phe(4-F)-PGWLVKNG was identified. This peptide demonstrated an EC50 value of 3.4 μM in a platelet-aggregation assay, which is 16-fold more potent than AYPGKF. Using this new protease-activated receptor 4 (PAR4) agonist peptide (AP), a platelet-rich plasma-aggregation assay using light-transmission aggregometry was developed and validated in a series of precision and reproducibility tests. PAR4 antagonist responses to PAR4 AP A-Phe(4-F)-PGWLVKNG (12.5 μM to 100 μM) were subsequently evaluated in this assay in vitro and ex vivo in a human study using BMS-986120, a PAR4 antagonist that entered clinical studies. 3632 GSYWYNVWF 0 Phage display (common panning) PMID:34079028 IgG Fc region human MDA-MB-231 cells and mouse EF43.fgf4 cells Not determined. Not determined. Ph.D.-12 phage display library (X12) ELISA peptide ligand binding to human IgGs 3633 NGFFEPWQVVYV 1 Phage display (competitive panning) 3 PMID:35007993 Maltose-binding protein (MBP) (MBP-VHH2,4-D) Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) ELISA The absorbance at 450 nm were measured by SpectraMax M5. The bound phages were competitively eluted by a 2,4-D standard solution and followed added to the wells only MBP coated to further remove undesired phages. A variable domain (VHH) to 2,4-dichlorophenoxyacetic acid (2,4-D) was used to isolated the peptidomimetic (NGFFEPWQVVYV) from phage display libraries using six panning conditions. Then the peptidomimetic and VHH were fused with the larger (LgN) and smaller piece (SmN) of split fragments of Nluc, respectively. In order to optimize the signal and sensitivity of the immunosensor, we explored the effects of the spacer between the peptidomimetic and LgN, the copy number of peptidomimetics, and the spacer between SmN and VHH, generating 24 combinations that allowed to conclude on their respective roles. Eventually, the developed “ready-to-use” immunosensor performed excellent signal-to-noise ratio and sensitivity, and could be applied to the detection of 2,4-D in real samples. 3634 SRHGQRALQALP(8/15)[1.1428±0.0442] 1 Phage display (common panning) 4 PMID:33567652 Anti-canine adenovirus 2 (CAdV-2) monoclonal antibody 2C1 Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) ELISA The analytical signal was obtained by measuring the absorbance at 450 nm. Data shown were reproduced from Figure 3a. The peptide, SRHGQRALQALP, is similar to the sequence "RIKQRETPAL" observed in the CAdV-2 Hex. 3635 SAPHSASDPHYS(9/15) SESAGGNHYSHP(2/15) FYQHSNPWAKYS(1/15) PLSRYKTRAYSC(1/15) VPESRYTVPSAT(1/15) PESXXDXXYS 5 Phage display (common panning) 4 PMID:33567652 Anti-canine adenovirus 2 (CAdV-2) monoclonal antibody 2C1 Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) ELISA The analytical signal was obtained by measuring the absorbance at 450 nm. Data shown were reproduced from Figure 3b. CAdV-2-specific neutralizing monoclonal antibodies The high affinity phage clones with mAb 7D7 displayed a consensus motif PESXXDXXYS (X is any aa), similar to the sequence "PESYKDRMYS" observed in the CAdV-2 Hex. 3636 CIHSPTALC(5124) CNAGHLSQC(4414) CNIKSSHVC(4296) CNMHTPMVC(4232) CISSSINHC(3804) CTLKNLALC(3787) CLTAKHMQC(3715) CLAYAHHTC(3457) CNWMINKEC(3267) CFATRADHC(3089) CDGLAKNSC(3028) CPKGDENTC(2982) CSAQQPASC(2966) CNDTKQGNC(2904) CGPTAKYIC(2867) CPSTVPWSC(2856) CVPSKPGLC(2826) CNSHTQGKC(2522) CSVGYDRNC(2487) CSENSPLLC(2478) CSKEATPFC(2445) CLDHSSKLC(2182) CNTSTMLHC(2181) CEQAHKLHC(2172) CTTKLPNSC(2066) CNANPWRLC(2054) CLNRYVADC(2027) 27 Phage display (common panning) PMID:35871689 Escherichia coli MSI001 Not determined. Not determined. Not determined. Ph.D.-C7C phage display library (CX7C) A representative heptapeptide CVTKLGSLC (VTK) was selected for fusion with the antibacterial peptide LL-37 to construct the specific-targeting antibacterial peptide VTK-LL37. We found that, in comparison with LL37, VTK-LL37 showed prominent bacteriostatic activity and an inhibitive effect on biofilm formation in vitro. In vivo experiments demonstrated that VTK-LL37 significantly inhibited bacterial growth, reduced HMGB1 expression, alleviated lesions of vital organs and improved the survival of mice subjected to CLP modeling. Furthermore, membrane DEGP and DEGQ were identified as VTK-binding proteins by proteomic methods. 3637 CVPKRRNNC[0.4316±0.0255] CLKRQRT[0.3551±0.0538] CRRRVS[0.3453±0.0353] CNRRRA[0.5789±0.0212] CRKRS[0.4203±0.027] CRKIR[0.3482±0.0324] 6 Phage display (subtractive panning) 4-5 PMID:35014500 Kidney injury molecule 1, KIM-1 Not determined. Not determined. Not determined. Byungheon Lee (Department of Biochemistry and Cell Biology, School Medicine, Kyungpook National University, Republic of Korea) ELISA Optical density data (450 nm) for each clone is expressed as the mean ± standard deviation (SD) of three independent experiments. Data shown were reproduced from Figure 1. Each biopanning round consisted of subtraction of phages nonspecifically bound to M-270 epoxy beads only and subsequent selection of phages bound to KIM-1 conjugated epoxy beads with 3 M ammonium sulfate, followed by amplification of the eluted phages. This peptide, with the sequence CNRRRA, not only showed a high imaging potential in vitro, allowing a strong detection of kidney injury molecule-1 (KIM-1) expressing cells by microscopy and flow cytometry but also generated a strong kidney-specific signal in live-imaging in vivo experiments in the context of a drug-induced kidney-injury mouse model. 3638 CTKTDVHFC[2.1512±0.0436] CIHSSTRAC[1.9349±0.0576] CMQTQRAHC[1.8843±0.0297] CTNANHYFC[1.1291±0.0279] CTYENHRTC[1.546±0.0645] CDPRHSKFC[0.9914±0.0436] CLAQSHPLC[1.3227±0.0297] 7 Phage display (competitive panning) 4 PMID:34635758 GTTYGVCSK-biotin and Domain III of the envelope (E) glycoprotein (DIII) Not determined. Not determined. Not determined. Ph.D.-C7C phage display library (CX7C) ELISA Optical density data (450 nm) for each clone is expressed as the mean ± standard deviation (SD). Data shown were reproduced from Figure 1B. The first round of panning: target: GTTYGVCSK-biotin (1 μg/well), phage elution: competitive elution with GTTYGVCSK-biotin (9 μg) in PBS (pH 7.2). The second round of panning: target: GTTYGVCSK-biotin (1 μg/well), phage elution: competitive elution with GTTYGVCSK-biotin (2 μg) in PBS (pH 7.2). The third round of panning: target: GTTYGVCSK-biotin (1 μg/well), phage elution: non-related peptide in CovaBuffer (pH 7.2) + 1 M glycine; followed by (ii) competitive elution with GTTYGVCSK-biotin (2 μg) in PBS (pH 7.2). The 4th round of panning: target: rDIII (0.3 μg/well), phage elution: 0.1% TBST (pH7.2) + 250 mM imidazole. Four cyclic peptides (CTKTDVHFC, CIHSSTRAC, CTYENHRTC, and CLAQSHPLC) showed significant blocking of the interaction between DIII and hBMECs, and ability to neutralize infection in cultured cells. 3639 SGVYKVAYDWQH[1.9855±0.0209] HYSWSWIAYSPG[3.0371±0.0139] 2 Phage display (competitive panning) 5 PMID:34635758 GTTYGVCSK-biotin and Domain III of the envelope (E) glycoprotein (DIII) Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) ELISA Optical density data (450 nm) for each clone is expressed as the mean ± standard deviation (SD). Data shown were reproduced from Figure 1B. The first round of panning: target: GTTYGVCSK-biotin (1 μg/well), phage elution: competitive elution with GTTYGVCSK-biotin (9 μg) in PBS (pH 7.2). The second round of panning: target: GTTYGVCSK-biotin (1 μg/well), phage elution: competitive elution with GTTYGVCSK-biotin (2 μg) in PBS (pH 7.2). The third round of panning: target: GTTYGVCSK-biotin (1 μg/well), phage elution: non-related peptide in CovaBuffer (pH 7.2) + 1 M glycine; followed by (ii) competitive elution with GTTYGVCSK-biotin (2 μg) in PBS (pH 7.2). The 4th round of panning: target: rDIII (0.3 μg/well), phage elution: 0.1% TBST (pH7.2) + 250 mM imidazole. The 5th round of panning: target: rDIII (0.3 μg/well), phage elution: competitive elution with GTTYGVCSK-biotin (2 μg) in PBS (pH 7.2). peptides, 4 cyclic peptides showed significant blocking of the interaction between DIII and hBMECs, and ability to neutralize infection in cultured cells. 3640 CSSTRESAC CRYSAARSC CRGFVVGRC CQRALMIAC 4 Phage display (in vivo) 3 PMID:34060472 Mouse mammary EF43.fgf4 cell Not determined. Not determined. Not determined. Ph.D.-C7C phage display library (CX7C) Binding assay Binding of individual phage clones to EF43.fgf4 cells was quantified by the counting of transducing units (TU) after host bacterial infection. The values of the relative TU were reproduced from Figure 1B and shown. Combinatorial phage display selections in vivo in tumor-bearing mice were performed as follows: animals received 10^9 TU iv of an unselected phage display random peptide library (displaying the insert CX7C). Tumors and control organs were collected after 24 hr of systemic circulation. For homing of individual phage clones in vivo, tumor-bearing mice were deeply anesthetized with 1–2% isofluorane and received 10^9 TU of targeted phage or insertless control phage, both administered iv side-by-side. Phage particles were recovered from tissue samples by bacterial infection and processed. We identified a cyclic peptide (CSSTRESAC) that specifically binds to a vitamin D receptor, protein disulfide-isomerase A3 (PDIA3) expressed on the cell surface of tumor-associated macrophages (TAM), and targets breast cancer in syngeneic triple-negative breast cancer (TNBC), non-TNBC xenograft, and transgenic mouse models. Systemic administration of CSSTRESAC to TNBC-bearing mice shifted the cytokine profile toward an antitumor immune response and delayed tumor growth. Moreover, CSSTRESAC enabled ligand-directed theranostic delivery to tumors and a mathematical model confirmed our experimental findings. 3641 ACSDRFNCPADEALCG(17) KCCNVINCCK(10) KCCTKNCDSTAHCT(9) RCDHCLCLQCS(8) TCCAFCVSCI(7) SCSTCNDSVSDCCL(6) QCMRQPCENCCR(6) TCSRTHQCTRPCCP(5) GCVSCCQGTCF(5) KCCTESPLCCT(3) SCPCEGCVNFSCF(2) SCHDCNTCGW(1) GCCCGSSVCH(1) RCPFPCTPQRCCY(1) RSCWRCSSAPFCCA(1) CVVLHSSEICDCS(1) DCCWQGWCICN(1) MCANCSEDPCCS(1) NCHPCNNCTNCK(1) QCRTQPSCPNCKCK(1) 20 Phage display (common panning) 5 PMID:33976148 Ice Not determined. Not determined. Not determined. XCXmCXnCXoCX phage library pool Our ice-affinity selection protocol enables the selection of a cyclic ice-binding peptide containing just 14 amino acids (KCCTKNCDSTAHCT). Mutational analysis identifies three residues, Asp8, Thr10 and Thr14, which are found to be essential for ice binding. 3642 NRPDSAQFWLHH[3.6] SITNGWKTVIRS[10.2] YIPEVRHSRAWH[16.7] HSSPHFSRHGLL[13.2] QPPLQPIPDALT[1.2] HPDIYFHPGNQR[42.9] GISWQQSHHLVA[34] ATWSHHLSSAGL[31.2] 8 Phage display (common panning) 3 PMID:36701953 Mucin-5B, MUC5B Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) Binding assay The amount of peptide in the supernatant of the control and test samples of each peptide was determined using capillary zone electrophoresis. The amount of peptide bound (shown as affinity value) was quantified by comparative analysis of the peak areas of control and test samples in % using BIOFOCUS Integrator software (Bio-Rad). The difference in the areas of peaks of control and test samples of each peptide corresponded to the amount of bound peptide. MBP-12 (HPDIYFHPGNQR) and MBP-14 (GISWQQSHHLVA) displayed the highest affinity to MUC5B. MBP-12 mildly stabilized the spinnbarkeit of serous saliva after overnight incubation and of mucous saliva at all timepoints tested. The addition of MBP-12 to a pellicle of unstimulated saliva on HA discs showed no additive protective effect against acid-induced demineralization. Epitope characterization suggested sulfo-Lewis(a) SO3–3Gal_1–3GlcNAc (galactose residue) as MBP-12 binding site on MUC5B. 3643 TLMVPRTGS(6) WCQVQSVCA(3) RWRRKALPW(2) GWRPKMVLR(2) RWRRKVLRH(2) LSSCFRFCS(1) WNMKFVWRS(1) RPRARVFRF(1) VWREPLHHH(1) ARGPRRGVR(1) RRVLSACVW(1) VTLRFNGWG(1) APAARNLEL(1) VLVPVGVVW(1) GAVSSHLDL(1) GISGVSGCS(1) LVYLVWDVW(1) TWRRRLLVH(1) TWRRRNLVH(1) TWRKRMRLL(1) GWRRKALMR(1) GWRRKVWLR(1) FWRRKVWHH(1) RWRRKVFLH(1) GWRRKVMRL(1) FWRRKVLHH(1) SWRRKIYHL(1) GWRRKLRVV(1) GWRRKSLWA(1) GWRKKSLRL(1) RWRKKAWHH(1) RWKRKVLEA(1) RWKRKVRVV(1) MWKRKVLVL(1) GWKRKTLSL(1) RWKRKSLGR(1) GWKRKYRQV(1) TWKRKWFHH(1) KYKRKRLFL(1) RWKKKSWLW(1) GRYRKNIMH(1) GWFRKMLFS(1) GWRPKTWIG(1) GWKPKWLLD(1) RWRAKVLPV(1) GWRVKSIPV(1) RWRVKSRKW(1) GWRTKVGVL(1) RWRPRVLPH(1) RWRPRNWRH(1) GWRARTLRV(1) RWRARVRRL(1) GWRARVLSL(1) GWRTRVLPL(1) RWRSRVLRI(1) RWRMRALFH(1) GWRPKRGYL(1) SWRPKRLPW(1) GWRAKRVIW(1) GWRAKRMMV(1) GWKAKRLSV(1) RWRVKRYSH(1) GWRVKRLVV(1) GWRLKRLVD(1) RWRVKRAML(1) GWRSKRLRV(1) GFRMKRLVV(1) GWRFKRLEL(1) GWRAKRRII(1) AWRPRRLVV(1) GWRARRLVI(1) GWRVRRLFT(1) GWKARKLVF(1) GWLAKRAHH(1) GWASKRLQV(1) RWFSKRLLH(1) VPRLKILVM(1) ASRLKVHHH(1) VVWGKMLSH(1) WGWLRFEHH(1) PGGWRRKALSV and PGGWRAKVLSV 80 Phage display (common panning) 7 PMID:34919980 Cathepsin G-like Not determined. Not determined. Not determined. X9 T7 phage display library A preference for a lysine in the P1 position of a substrate, arginines in positions P2 and P3, and the aromatic amino acid tryptophane in the P4 position was observed. Based on the sequence alignment we could identify a consensus sequence for this protease as being PGGWRRK↓ALSV. Mass spectrometry analysis of a peptide with the consensus sequence obtained by phage display showed that cleavage of this peptide occurred after the conserved Lys (K) residue. 3644 WDLPPIGRLSGN(50.0%) SWMPPILRSPAV(26.3%) THLPPIMRNLQF(5.7%) SWLPNIQRHWLS(2.6%) HLPPILRMLDLV(2.6%) HLPPIQRTPTYA(2.6%) LPPIVRLPGLLH(2.6%) FPFGPINRDMTA(2.6%) NGVWLPPIARVL(2.6%) LPPIXRX 9 Phage display (common panning) 3 PMID:36312416 Good Vibes phage, GV Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) ELISA The absorbance was measured at 450 nm using an Infinite 200 Rx plate reader. The heat map of Figure 3C shows the cross-reactivity of GV monoclonal phages against GV (target), MAT (same family as GV), cowpea mosaic virus (CPMV, unrelated plant virus), and bovine serum albumin (BSA). We screened a phage display library for peptides that bind to the Good Vibes phage (GV), which lyses the bacterial pathogen Pseudomonas aeruginosa. Isolated monoclonal library phages featured a highly conserved consensus motif, LPPIXRX. The corresponding peptide WDLPPIGRLSGN was synthesized with a GGGSK linker and conjugated to cyanine 5 or biotin. The specific binding of the LPPIXRX motif to GV in vitro was confirmed using an enzyme-linked immunosorbent assay. We demonstrated imaging and tracking of GV in bacterial populations using the fluorescent targeting peptide and flow cytometry. 3645 RWKDTAYALTNN(4)[NB] SWHWHTHVRHQM(3)[0.04 ± 0.01] WGHSHFSHWKGR(3)[6.86 ± 2.63] HHLRIPYALDQT(2)[NB] SHRWQVWSRDRA(2)[NB] ASANDNRLRYTY(2)[NB] LDRPSSLAHLAS(2)[2.14 ± 0.69] RHHSSNPRDTAP(2)[8.82 ± 0.01] 8 Phage display (common panning) 5 PMID:36195696 Cytotoxic T-lymphocyte protein 4 T-lymphocyte activation antigen CD80 Not determined. Not determined. Ph.D.-12 phage display library (X12) The human CTLA4 binding peptides to hCTLA-4 protein affinity was measured by microscale thermophoresis (MST). The dissociation constant (Kd, μM) was determined and shown. The LC4 peptide (WGHSHFSHWKGR) was modified to improve its tumour-targeting ability and reduce peripheral immune system activation, which could block the CTLA-4/CD80 interaction. The LC4 peptide as a result, like other immune checkpoint inhibitors (ICIs), exerts anti-tumour effects by refreshing T cell function, and also activates the peripheral immune system. We used the PLGLAG peptide as a linker at the C-terminal of LC4 to connect with a tumourtargeting peptide RGD to increase the tumour tissue targeting ability, and obtain LC4-PLG-RGD. Further experiments demonstrated that the anti-tumour LC4-PLG-RGD activity was better than LC4 in vivo, and the ability to activate the peripheral immune system was weakened. 3646 ITNAPIKDLTP NSNDFRPGGPET SNKNLDTRILTK 3 Phage display (common panning) 3 PMID:34260404 APTYSW peptide CD81 antigen Not determined. Not determined. Ph.D.-12 phage display library (X12) 3647 LPLSTQH 1 Phage display (common panning) 5 PMID:34673862 Fe3O4 Not determined. Not determined. Not determined. Ph.D.-7 phage display library (X7) Special M13 with Fe3O4 affinity pIII-peptide (FAP-M13) was biopanned for strongly binding towards bare Fe3O4 with the “hook”-like pIII-peptide (N-LPLSTQH-C). TEM observation confirmed the direct grasp of FAP-M13 on bare Fe3O4, forming the magnetic (FAP-M13)-Fe3O4 virus hydrogel. 3648 CRTLPFHEC(7/80)[5.9 ± 0.2] CEKMVATHC(4/80)[not determined] CRTLPWNQC(3/80)[2.3 ± 0.6] CRTIPFTHC(2/80)[not tested] CRTMEYTSC(2/80)[not tested] CRTLPYHLC(2/80)[2.7 ± 0.2] CRTQPYNQC(1/80)[not tested] 7 Phage display (subtractive panning) 5 PMID:33668971 WD repeat-containing protein 5 Not determined. Not determined. Not determined. Ph.D.-C7C phage display library (CX7C) The fluorescence polarization was measured in Synergy™ Neo2 Multi-Mode Microplate Reader using an excitation filter at 485 nm and an emission filter at 535 nm. The EC50 (μM) value was determined and shown. To remove false-positive target-unrelated peptides (TUPs) which directly bind to Ni-NTA magnetic beads, the library was depleted 2 times by incubating the phage library directly with Ni-NTA magnetic beads before the biopanning against the target in the second and third rounds. The detailed binding interactions were revealed by solving a WDR5-peptide cocrystal structure. 3649 WSLGYTG(8.9%) 1 Phage display (common panning) 3 PMID:35328728 T-cell surface glycoprotein CD4 Not determined. Not determined. Not determined. Ph.D.-7 phage display library (X7) We report the isolation of a fast-propagating white clone (displaying WSLGYTG peptide) identified through screening against a recombinant protein. Sanger sequencing demonstrated that white plaques are not contamination from environmental M13-like phages, but derive from the library itself. Whole genome sequencing revealed that the white color of the plaques results from a large 827-nucleotide genomic deletion. The phenotypic characterization of propagation capacity through plaque count- and NGS-based competitive propagation assay supported the higher propagation rate of Ph-WSLGYTG clone compared with the library. 3650 GLHTSATNLYLH[0.2704±0.0069] DSQFNKYSIATV[0.2676±0.0194] MHPNAGHGSLMR[0.2055±0.0076] DEVPPGMTLPRP[0.2338±0.0262] SGVYKVAYDWQH[0.2262±0.0111] 5 Phage display (common panning) 3 PMID:33121397 Envelope protein 2, E2 Not determined. Not determined. Not determined. Ph.D.-12 phage display library (X12) ELISA The absorbance was measured at 450 nm using an ELISA plate reader. ELISA values were reproduced from Figure 3 and shown. Five peptides were identified as potential binders based on their robust reactivity to the bait protein. The selected peptides appeared to interact with the crucial residues that were notably exposed on the surface of E1-E2 trimeric structure.