Discovery of exolytic heparinases and their catalytic mechanism and potential application

Qingdong Zhang,Fuchuan Li,Min Yuan,Xiu-Lan Chen,Peng Wang,Xiangxue Chen,Lianli Chi,Yu-Zhong Zhang,Lin Wei,Deling Shi,Danrong Lu,Min Du,Hai-Yan Cao

doi:10.1038/s41467-021-21441-8

Abstract

Heparinases (Hepases) are critical tools for the studies of highly heterogeneous heparin (HP)/heparan sulfate (HS). However, exolytic heparinases urgently needed for the sequencing of HP/HS chains remain undiscovered. Herein, a type of exolytic heparinases (exoHepases) is identified from the genomes of different bacteria. These exoHepases share almost no homology with known Hepases and prefer to digest HP rather than HS chains by sequentially releasing unsaturated disaccharides from their reducing ends. The structural study of an exoHepase (BIexoHep) shows that an N-terminal conserved DUF4962 superfamily domain is essential to the enzyme activities of these exoHepases, which is involved in the formation of a unique L-shaped catalytic cavity controlling the sequential digestion of substrates through electrostatic interactions. Further, several HP octasaccharides have been preliminarily sequenced by using BIexoHep. Overall, this study fills the research gap of exoHepases and provides urgently needed tools for the structural and functional studies of HP/HS chains.

Highlights

Heparinases (Hepases) are critical tools for the studies of highly heterogeneous heparin (HP)/heparan sulfate (HS)
Carbohydrate-Active Enzyme database and Simple Modular Architecture Research Tool analyses showed that the BIexoHep protein is composed of three domains: an N-terminal signal peptide, a DUF4962 superfamily domain and a C-terminal Hepase II/III superfamily domain
The discovery of the Hepase BIexoHep with exolytic activity led to the identification of a Hepase family, PL15_2, a subfamily of the PL15 family that has been known as the assembly of exolytic alginate lyases[31], which is different from Hepases I, II, and III, belong to the PL13, PL21, and PL12 families[16], respectively

Summary

Introduction

Heparinases (Hepases) are critical tools for the studies of highly heterogeneous heparin (HP)/heparan sulfate (HS). The various structural characteristics of HP/HS permit these molecules to interact with various proteins, such as enzyme inhibitors[3], chemokines[4], growth factors[5], morphogens[6], and other signaling proteins, to participate in various physiological and pathological processes, such as coagulation[7], cell adhesion[8], inflammation[4], cell migration[3], differentiation[9], and even pathogenic infections[10]. These vital biological roles have attracted great attention to the structural and functional studies and the clinical applications of HP/. These modifications can result in numerous disaccharide variations in HP/HS polysaccharides, which make HP/HS the most sophisticated polymers in nature[13]

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Results

Conclusion