Switchable Enzymatic Accessibility For Precision Cell-Selective Surface Glycan Remodeling

As one of the basic components of cells,glycans are the most complex and diverse biological macromolecules in nature.They serve as a critical handle for mediating cell recognition,cell communication,cell adhesion,and immune response.Glycosylation is a non-template-driven biosynthesis process that is inherently cell type or cell population-dependent.Aberrant glycosylation is implicated in disease(e.g.,cancer)progression.The ability to target a specific type or population of cells for glycan remodeling is therefore essential for the full understanding and precise modulation of glycan functions.Cell-selective glycan manipulation can also provide a potential tool for tissue-specific glycan imaging and immune therapy.Previous attempts at developing cell-selective glycan remodeling methods rely on either indirect internal metabolic pathway or direct external enzymatic action.The internal metabolic approach allows the achievement of cell specificity as it offers a division of cell-targeting metabolic precursor delivery stage and on-cell glycan remodeling stage,but it can suffer from undesired whole-cell perturbation(on both intracellular and extracellular proteins),unintended glycosylation site,and lengthy protocol duration issues.The documented external enzymatic approach is plagued by compromised cell selectivity because the enzyme is catalytically active toward unavoidable glycan remodeling on non-targeted cell surface during its supposedly guided delivery to target cell surface.Hence,the stringent control of enzymatic catalytic activity at the cell-targeting delivery stage and on-cell glycan remodeling stage is essential for cell-selective glycan remodeling.At present,the enzymatic activity is mainly regulated by modifing the chemical molecule which can be cleaved(e.g.,photolysis)at the specific site of the enzyme,so that the enzyme is biologically inert.Another method is to covalently immobilize enzyme in nanostructure to prevent its contact with interacting molecules.These caging methods typically target the off-on switchability of intrinsic enzyme function,and the covalent can lead to slow restoration kinetics and even partial loss of catalytic activity.Furthermore,each enzyme that is intended to be caged demands a new design and synthesis of the caged molecules,which is tedious and not universal.In recent years,the research on metal-organic framework(MOF)has developed rapidly in the interdisciplinary fields including basic chemistry,nanotechnology and biomedicine.As the most representative subclass of MOF,ZIF-8 is formed through the coordination between 2-methyl imidazole(2-MIM)and zinc ions,and can be rapidly degraded in the slightly acidic environment.ZIF-8 has good water stability and biocompatibility,and is widely used in sensing,drug carrier,immobilized enzyme and other fields.To address these issues,herein we report a switchable enzymatic accessibility(SEA)method for precision cell-selective surface glycan remodeling.SEA is a general,versatile designer platform for cell-selective surface structural remodeling,and exploits the combined use of an enzymatic accessibility-regulating caging probe component(ZIF-8)and a cell-targeting recognition probe component(Apt).Firstly,the probe is synthesized by biomimetic mineralization:galactose oxidase(GO)was encapsulated in the cavities of ZIF-8 to form GO@ZIF-8 complex.Afterwards,aptamer is coupled through electrostatic adsorption to obtain the final probe.Briefly,a two-stage protocol is used:the first stage is the delivery of ZIF-8-encapsulated,catalytically inaccessible enzyme to target cell surface via aptamer binding(the accessibility of enzyme is blocked by ZIF-8 and no glycan remodeling occurs at this stage;if the enzyme is not blocked and glycan remodeling occurs at this stage,non-targeted cells will be modified);the second stage is the local release of enzyme(switched to the catalytically accessible state)on target cell surface via MOF disassembly(local release is possible due to slow diffusion rate for a macromolecular-sized species such as enzyme)for glycan remodeling of only target cells.This strategy makes the use of accessibility-sifting property of ZIF-8(with respect to substrate size dimension)and achieve cell-selective glycan remodeling.The SEA protocol features a modular and generically adaptable design,a very short protocol duration(?30 min or even shorter),and a very high spatial resolving power(ability to differentiate immediately neighboring cell lines).