Carbohydrate-protein interactions by "clicked" carbohydrate self-assembled monolayers.
A Huisgen 1,3-dipolar cycloaddition "click chemistry" was employed to immobilize azido sugars (mannose, lactose, alpha-Gal) to fabricate carbohydrate self-assembled monolayers (SAMs) on gold. This fabrication was based on preformed SAM templates incorporated with alkyne terminal groups, which could further anchor the azido sugars to form well-packed, stable, and rigid sugar SAMs. The clicked mannose, lactose, and alpha-Gal trisaccharide SAMs were used in the analysis of specific carbohydrate-protein interactions (i.e., mannose-Con A; ECL-lactose, alpha-Gal-anti-Gal). The apparent affinity constant of Con A binding to mannose was (8.7 +/- 2.8) x 10(5) and (3.9 +/- 0.2) x 10(6) M(-1) measured by QCM and SPR, respectively. The apparent affinity constants of lactose binding with ECL and alpha-Gal binding with polyclonal anti-Gal antibody were determined to be (4.6 +/- 2.4) x 10(6) and (6.7 +/- 3.3) x 10(6) M(-1), respectively by QCM. SPR, QCM, AFM, and electrochemistry studies confirmed that the carbohydrate SAM sensors maintained the specificity to their corresponding lectins and nonspecific adsorption on the clicked carbohydrate surface was negligible. This study showed that the clicked carbohydrate SAMs in concert with nonlabel QCM or SPR offered a potent platform for high-throughput characterization of carbohydrate-protein interactions. Such a combination should complement other methods such as ITC and ELISA in a favorable manner and provide insightful knowledge for the corresponding complex glycobiological processes.
Department of Biochemistry and Chemistry, The Ohio State University, Columbus, Ohio 43210, USA.
Yun
Y
Zhang
Sanzhong
S
Luo
Yijun
Y
Tang
Lei
L
Yu
Kuang-Yu
KY
Hou
Jin-Pei
JP
Cheng
Xiangqun
X
Zeng
Peng George
PG
Wang
D002241:Carbohydrates / Q000737:chemistry*; D020650:Combinatorial Chemistry Techniques; D004563:Electrochemistry; D004566:Electrodes; D008567:Membranes, Artificial; D018625:Microscopy, Atomic Force; D015394:Molecular Structure; D011506:Proteins / Q000737:chemistry*; D012680:Sensitivity and Specificity; D020349:Surface Plasmon Resonance; D013997:Time Factors
D002241:Carbohydrates; D008567:Membranes, Artificial; D011506:Proteins