Figure S1. Expression efficiency of rd2bpl3 for various expression hosts. (A) BL21(DE3), (B)
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1 1 Supplementary Figure S Figure S1. Expression efficiency of rd2bpl3 for various expression hosts. (A) BL21(DE3), (B) BL21(DE3)pLysS, (C) BL21-CodonPlus(DE3)-RIL, (D) Rosetta(DE3). M, Molecular weight marker; Lane 1, un-induced lysate; Lane 2, IPTG induced insoluble fraction; Lane 3, IPTG-induced soluble fraction. Arrows indicate target proteins. 9
2 10 Supplementary Figure S Figure S2. Confirmation of the peptide sequence using LC-MS/MS. (A) Mass spectra of trypsin- digested rd2bpl3; (B) Peptide sequence and matching with mass spectrum data.
3 14 Supplementary Figure S Figure S3. Purification of native BPL3. M, Molecular weight marker; Lane 1, Purified BPL3. Arrows indicate purified proteins. 19
4 20 Supplementary Figure S Figure S4. Predicted 3-dimensional structure of BPL3. Structures were analyzed by protein sequence similarity analysis using I-TASSER program ( and Swiss-Prot 3D structure prediction program ( with normal parameters; C-score was typically in the range of [-5, 2], where a C-score of a higher value signifies a model with a higher confidence and vice-versa; (A) native BPL3, c-score was 0.07; (B) enlarged image of native BPL3 binding sites; (C) rd2bpl3, c-score was -3.31; (D) enlarged image for rd2bpl3 binding sites; (E) predicted 3D structure was calculated using Swiss-Model. BPL3 was matched with Helix Pomatia agglutinin (HPA) 30
5 31 32 Table S1. Glycan array of recombinant and native BPL3. Binding signals were normalized using a program provided by RayBioTech Substrates Positive Control Glycan Structures - RFU (Normalized) rd2bpl3 Native BPL3 29,419 29,419 Neg β-glc-sp 860± ±52 2 β-gal-sp 510± ±44 3 α-man-sp 1,322±366 1,025±57 4 α-fuc-sp 1,313± ±72 5 α-rha-sp 86±17 101±25 6 β-glcnac-sp 3,164± ±112 7 β-galnac-sp 698±545 97±15 8 Tobramycin 492±13 121±41 9 Gal-β-1,3-GlcNAc-β-Sp 1,646± ±65 10 Gal-α-1,3-Gal-β-1,3-GlcNAc-β-Sp 729± ± Neu5Ac-α-2,3-Gal-β-1,3-GlcNAc-β-Sp 165±28 120±55 12 Neu5Ac-α-2,6-Gal-β-1,3-GlcNAc-β-Sp 183± ±44 13 Neu5Gc-α-2,3-Gal-β-1,3-GlcNAc-β-Sp 325± ±23 14 Neu5Gc-α-2,6-Gal-β-1,3-GlcNAc-β-Sp 229± ±33 15 Gal-β-1,3-(Fuc-α-1,4)-GlcNAc-β-[Lewis A] Sp 660±290 70±23 16 Gal-β-1,4-Glc-β-Sp 204± ±14 17 Gal-α-1,3-Gal-β-1,4-Glc-β-Sp 644± ±23 18 Gal-α-1,4-Gal-β-1,4-Glc-β-Sp 187±106 85±9 19 GlcNAc-β-1,3-Gal-β-1,4-Glc-β-Sp 786± ±84 20 GalNAc-β-1,3-Gal-β-1,4-Glc-β-Sp 87±32 58±19 21 Neu5Ac-α-2,3-Gal-β-1,4-Glc-β-Sp 46±13 11±21 22 Neu5Ac-α-2,6-Gal-β-1,4-Glc-β-Sp 57±15 7±32 23 Neu5Gc-α-2,3-Gal-β-1,4-Glc-β-Sp 266±160-36±72 24 Neu5Ac-α-2,6-Gal-β-1,4-Glc-β-Sp 991± ±24 25 Gal-β-1,4-(Fuc-α-1,3)-Glc-β-Sp 647±51 255±42 26 GalNAc-β-1,3-Gal-α-1,4-Gal-β-1,4-Glc-β-Sp 132±16 130±10 27 GlcNAc-β-1,6-GlcNAc-β-Sp 864± ± P-GlcNAc-β-1,4-Man-β-Sp 148±77 116±23 29 Glc-α-1,2-Gal-α-1,3-Glc-α-Sp 260±157 78±13 30 Gal-β-1,3-GalNAc-α-Sp 64±13 5,075± Gal-β-1,4-GlcNAc-β-Sp 66±38 105±20 32 Gal-β-1,4 -(Fuc-α-1,3)-GlcNAc-β-[Lewis X] Sp 93±29 104± Neu5Ac-α-2,3-Gal-β-1,4-(Fuc-α-1,3)-GlcNAc-β- [Sialyl Lewis X]-Sp 235±130 25±7 Neu5Ac-α-2,3-Gal-β-1,3-(Fuc-α-1,4)-GlcNAc-β- [Sialyl Lewis A]-Sp 973± ±32 Neu5Gc-α-2,3-Gal-β-1,3-(Fuc-α-1,4)-GlcNAc-β- [Sialyl Lewis A]-Sp 346±115 45± Gal-α-1,4-Gal-β-1,3-GlcNAc-β-Sp 721±328 1,082± Gal-β-1,4-GlcNAc-β-1,3-Gal-β-1,4-Glc-β- [LNnT]-Sp 797± ± GlcA-β-1,4-GlcNAc-α-1,4-GlcA-β-Sp 401± ±23 39 GlcNAc-β-1,6-(Gal-β-1,3)-GalNAc-α-O-Ser- Sp4 77±48 1,398± Neu5Ac-α-2,3Gal-β-1,4-(6S)GlcNAc-β-Sp 326±222 72±13 41 GalNAc-β-1,4-GlcNAc-β-Sp2 993± ±45 42 Neu5Ac-α-2,8-Neu5Ac-α-2,3-Gal β-1,4-glc-β-sp 102±79 65±16 43 Neu5Gc-α-2,8-Neu5Ac-α-2,3-Gal-β-1,4-Glc-β-Sp 40±21 46±13 44 GalNAc-α-1,3-(Fuc-α-1,2)-Gal-β-1,4-Glc-β- [Blood A antigen tetrose]-sp1 70±18 256±48
6 45 GlcNAc-β-1,2-Man-α-Sp 1,290± ±53 46 Neu5Ac-α-2,3-Gal-β-Sp1 565±90 42±4 47 Gal-β-1,3 -GalNAc-β-1,3-Gal-β-Sp1 53±34 12±29 48 Glc-α-1,2-Gal-α-Sp 264±150 74±28 49 Gal-β-1,4-(Fuc-α-1,3)-GlcNAc-β-1,3-Gal-β-Sp1 43±39 2±25 Neu5Ac-α-2,3-Gal-β-1,4-(Fuc-α-1,3)-Glc-β- [3- Sialyl-3-fucosyllactose/ F-SL]-Sp1 54±9 2± GlcNAc-β-1,4-GlcNAc-β-Sp1 135±3 57±34 52 β-d-glca-sp 595±257 99±10 53 Gal-β-1,4-(6S)GlcNAc-β-Sp 169±71 84±16 54 GlcNAc-α-1,3-(Glc-α-1,2-Glc-α-1,2)-Gal-α-1,3-Glcα-Sp 214±90 64±24 55 Gal-β-1,3-GalNAc-β-1,4-(Neu5Gc-α-2,3)-Gal-β-1,4- Glc-β-Sp1 138±37 69±8 56 Sisomicin Sulfate 105±9 80±24 57 GalNAc-α-1,3-(Fuc-α-1,2)-Gal-β-[Blood A antigen trisaccharide]-sp1 59±30 181±73 58 Fuc-α-1,2-Gal-β-1,4-GlcNAc-β-[Blood H antigen trisaccharide]-sp1 57±21 72±5 59 Gal-α-1,3-(Fuc-α-1,2)-Gal-β-[Blood B antigen trisaccharide]-sp1 158±92 72±24 60 Fuc-α-1,2-Gal-β-1,3-GlcNAc-β-1,3-Gal-β-1,4-Glc-β- [LNFP I]-Sp1 81±51 44±17 61 Fuc-α-1,2-Gal-β-1,4-Glc-β-[Blood H antigen trisaccharide]-sp1 148±60 84±24 62 Gal-α-1,3-(Fuc-α-1,2)-Gal-β-1,4-Glc-β-[Blood B antigen tetrasaccharide]-sp1 108±20 14±7 63 (Fuc-α-1,2)-Gal-β-1,4-(Fuc-α-1,3)-GlcNAc-β- [ Lewis Y]-Sp1 49±28 24±20 64 (Fuc-α-1,2)-Gal-β-1,3-(Fuc-α-1,4)-GlcNAc-β- [ Lewis B]-Sp1 122±31 60±18 Gal-β-1,3-(Fuc-α-1,4)-GlcNAc-β-1,3-Gal-β-1,4-(Fucα-1,4)-Glc-β- [Lewis A]-Sp1 45±10 47± Gal-β-1,3-GalNAc-β-Sp1 39±16 17±9 67 Gal-β-1,3-(Neu5Ac-α-2,6)-GalNAc-β-Sp 318±43 240±32 68 Neu5Ac-α-2,6-Gal-β-1,3-GalNAc-β-Sp 247±59 101± Neu5Ac-α-2,6-Gal-β-1,3-(Neu5Ac-α-2,6)-GalNAc-β- Sp 421±59 2,518±904 Neu5Ac-α-2,3-Gal-β-1,3-(Neu5Ac-α-2,6)-GalNAc-β- Sp 789± ±12 Neu5Ac-α-2,6-(Neu5Ac-α-2,3)-Gal-β-1,3-GalNAc-β- Sp 993±349 96±24 GalNAc-β-1,4-(Neu5Ac-α-2,3)-Gal-β-1,4-Glc-β- [GM2]-Sp 191±44 338±96 GalNAc-β-1,4-(Neu5Ac-α-2,8-Neu5Ac-α-2,3)-Gal-β- 1,4-Glc-β-[GD2]-Sp 131±66 154± Gal-α-1,4-Gal-β-1,4-GlcNAc-β-Sp1 238±289 12±22 75 β-d-rha-sp 615±331 99±8 76 Glc-α-1,4-Glc-β-Sp1 227±59 265±19 77 Glc-α-1,6-Glc-α-1,4-Glc-β-Sp1 941± ±57 78 Maltotriose-β-Sp1 796±340 19±18 79 Glc-α-1,6-Glc-α-1,6-Glc-β-Sp1 376±148 62±38 80 Maltotetraose-β-Sp1 181±89 43±45 81 GlcNAc-α-1,4-GlcA-β-1,4-GlcNAc-α1,4-GlcA-β-Sp 200± ±61 82 Maltohexaose-β-Sp1 108±54 47±24 83 Maltoheptaose-β-Sp1 72±20 90±17 84 Acarbose-β-Sp1 113±53 33±14
7 33 85 D-Pentamannuronic acid-β-sp1 150±41 146±17 86 L-Pentaguluronic acid-β-sp1 529±439 89±7 87 D-Cellose-β-Sp1 1,119± ±69 88 Gal-α-1,3-Gal-β-Sp1 368± ±20 89 β-1,4-xylotetrose-sp1 155±110 84±9 90 Chitin-Trisaccharide-Sp1 45±31 44±28 91 KDN-α-2,8-Neu5Ac-α-2,3-Gal-β-1,4-Glc-β-Sp 108±20 59±16 92 Neu5Ac-α-2,8-Neu5Gc-α-2,3-Gal-β-1,4-Glc-β-Sp 177±127 34±15 93 Neu5Ac-α-2,8-Neu5Ac-α-2,8-Neu5Ac-α-2,3-Gal-β- 1,4-Glc-β-Sp3 122±53 72±24 94 Neu5Ac-α-2,8-Neu5Ac-α-2,6-Gal-β-1,4-Glc-Sp5 102±42 35±18 Gal-β-1,3-GalNAc-β-1,4-(Neu5Ac-α-2,3)-Gal-β-1,4-95 Glc-β-Sp1 115±26 88±28 96 Gentamicin Sulfate 104±25 76±19 97 Kanamycin Sulfate 128±44 102±6 98 Geneticin Disulfate Salt (G418) 61±19 28±23 99 Neomycin Trisulfate 333±36 129± SGP 75±6 52±22 34
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