Revealing the Mechanisms of Protein Disorder and N-Glycosylation in CD44-Hyaluronan Binding Using Molecular Simulation

Abstract

The extracellular N-terminal hyaluronan binding domain (HABD) of CD44 is a small globular domain that confers hyaluronan (HA) binding functionality to this large transmembrane glycoprotein. When recombinantly expressed by itself, HABD exists as a globular water-soluble protein that retains the capacity to bind HA. This has enabled atomic-resolution structural biology experiments that have revealed the structure of HABD and its binding mode with oligomeric HA. Such experiments have also pointed to an order-to-disorder transition in HABD that is associated with HA binding. However, it had remained unclear how this structural transition was involved in binding since it occurs in a region of HABD distant from the HA-binding site. Furthermore, HABD is known to be N-glycosylated, and such glycosylation can diminish HA binding when the associated N-glycans are capped with sialic acid residues. The intrinsic flexibility of disordered proteins and of N-glycans makes it difficult to apply experimental structural biology approaches to probe the molecular mechanisms of how the order-to-disorder transition and N-glycosylation can modulate HA binding by HABD. We review recent results from molecular dynamics simulations that provide atomic-resolution mechanistic understanding of such modulation to help bridge gaps between existing experimental binding and structural biology data. Findings from these simulations include: Tyr42 may function as a molecular switch that converts the HA-binding site from a low affinity to a high affinity state; in the partially disordered form of HABD, basic amino acids in the C-terminal region can gain sufficient mobility to form direct contacts with bound HA to further stabilize binding; and terminal sialic acids on covalently attached N-glycans can form charge-paired hydrogen bonding interactions with basic amino acids that could otherwise bind to HA, thereby blocking HA binding to glycosylated CD44 HABD.

The association between plasma hyaluronan level and plaque types in ST-Segment–Elevation Myocardial Infarction patients

“…In reference with a previous study, we deduced that plasma HA binds to the N-terminal hyaluronan binding domain (HABD) of CD44 in an in ammatory state to consume free CD44 in plasma. 26 Although the mRNA expression of CD44v1 and CD44v6 in PBMCs has been reported to be different between plaque rupture and erosion in the past, 7 there was no difference in soluble CD44 between plaque erosion and rupture in our study. Soluble CD44 is not only regulated by HA but also other factors, such as cytokines and shedding from immune cells.…”

Efficient Construction of Atomic-Resolution Models of Non-Sulfated Chondroitin Glycosaminoglycan Using Molecular Dynamics Data

“…Additionally, some studies have used results from LC-MS [45], X-ray crystallography [46], and NMR [46][47][48][49][50][51] to compare and validate conformational data from molecular dynamics (MD) simulations. This suggests that MD simulations can produce results complementary to experimental analysis methods by providing realistic three-dimensional atomic-resolution molecular models of GAG conformational ensembles [52][53][54][55][56]. Compact non-sulfated chondroitin 20-mer conformation arising from flexible glycosidic linkages (red) between monosaccharide rings (GalNAc in blue and GlcA in cyan).…”

Molecular mechanism for bidirectional regulation of CD44 for lipid raft affiliation by palmitoylations and PIP2

“…The Cluster-of-Differentiation-44 protein (CD44) is a versatile molecule that is involved in a variety of cellular processes, including inflammation, hematopoiesis, cell migration and cancer invasiveness [1][2][3]. The protein consists of an ectodomain (ED), a single transmembrane domain (TMD), and a cytoplasmic tail (CT).…”