8W4L

Crystal structure of closed conformation of human immunoglobulin Fc in presence of EndoSz

Summary for 8W4L

• Entry DOI: 10.2210/pdb8w4l/pdb
• Related: 8W4G
• Descriptor: Immunoglobulin gamma-1 heavy chain, N-acetyl-alpha-neuraminic acid-(2-6)-beta-D-galactopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-2)-alpha-D-mannopyranose-(1-6)-[2-acetamido-2-deoxy-beta-D-glucopyranose-(1-2)-alpha-D-mannopyranose-(1-3)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta-D-glucopyranose, beta-D-galactopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-2)-alpha-D-mannopyranose-(1-3)-[beta-D-galactopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-2)-alpha-D-mannopyranose-(1-6)]beta-D-mannopyranose-(1-4)-2-acetamido-2-deoxy-beta-D-glucopyranose-(1-4)-[alpha-L-fucopyranose-(1-6)]2-acetamido-2-deoxy-beta-D-glucopyranose, ... (4 entities in total)
• Functional Keywords: n-linked glycan, glycoside hydrolase, transglycosylation, immune system
• Biological source: Homo sapiens (human)
• Total number of polymer chains: 2
• Total formula weight: 54034.27

Authors

Guan, H.H.,Lin, C.C.,Hsieh, Y.C.,Chen, C.J. (deposition date: 2023-08-24, release date: 2024-07-03, Last modification date: 2024-10-30)

Primary citation

Hsieh, Y.C.,Guan, H.H.,Lin, C.C.,Huang, T.Y.,Chuankhayan, P.,Chen, N.C.,Wang, N.H.,Hu, P.L.,Tsai, Y.C.,Huang, Y.C.,Yoshimura, M.,Lin, P.J.,Hsieh, Y.H.,Chen, C.J.
Structure-Based High-Efficiency Homogeneous Antibody Platform by Endoglycosidase Sz Provides Insights into Its Transglycosylation Mechanism.
Jacs Au, 4:2130-2150, 2024

PubMed Abstract: Monoclonal antibodies (mAbs) have gradually dominated the drug markets for various diseases. Improvement of the therapeutic activities of mAbs has become a critical issue in the pharmaceutical industry. A novel endo-β--acetylglucosaminidase, EndoSz, from is discovered and applied to enhance the activities of mAbs. Our studies demonstrate that the mutant EndoSz-D234M possesses an excellent transglycosylation activity to generate diverse glycoconjugates on mAbs. We prove that EndoSz-D234M can be applied to various marketed therapeutic antibodies and those in development for antibody remodeling. The remodeled homogeneous antibodies (mAb-G2S2) produced by EndoSz-D234M increase the relative ADCC activities by 3-26-fold. We further report the high-resolution crystal structures of EndoSz-D234M in the -form at 2.15 Å and the complex form with a bound G2S2-oxazoline intermediate at 2.25 Å. A novel pH-jump method was utilized to obtain the complex structure with a high resolution. The detailed interactions of EndoSz-D234M and the carried G2S2-oxazoline are hence delineated. The oxazoline sits in a hole, named the oxa-hole, which stabilizes the G2S2-oxazoline in transit and catalyzes the further transglycosylation reaction while targeting Asn-GlcNAc (+1) of Fc. In the oxa-hole, the H-bonding network involved with oxazoline dominates the transglycosylation activity. A mobile loop2 (a.a. 152-159) of EndoSz-D234M reshapes the binding grooves for the accommodation of G2S2-oxazoline upon binding, at which Trp154 forms a hydrogen bond with Man (-2). The long loop4 (a.a. 236-248) followed by helix3 is capable of dominating the substrate selectivity of EndoSz-D234M. In addition, the stepwise transglycosylation behavior of EndoSz-D234M is elucidated. Based on the high-resolution structures of the -form and the bound form with G2S2-oxazoline as well as a systematic mutagenesis study of the relative transglycosylation activity, the transglycosylation mechanism of EndoSz-D234M is revealed.

PubMed: 38938812
DOI: 10.1021/jacsau.4c00004

Experimental method

X-RAY DIFFRACTION (3.1 Å)