The Polysialyltransferases Interact with Sequences in Two Domains of the Neural Cell Adhesion Molecule to Allow Its Polysialylation

Thompson, M.; Foley, Deirdre; Colley, Karen J.

doi:10.1074/jbc.m112.438374

Cited by 16 publications

(19 citation statements)

References 44 publications

(62 reference statements)

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“…4d) in proximity to Arg82, which has also been shown to contribute to NCAM binding 49 . Consistently with this orientation, the Ig5 domain of the nonpolysialylated homolog, the olfactory cell adhesion molecule, contains an Nglycan in the region close to the PBR domain, which, when engineered into NCAM, abro gated polysialylation 55 .…”

Section: A Model Of Polysialyltransferase-acceptor Interactionsmentioning

confidence: 54%

“…The proximity of Asn74 to NCAM Ig5 in our model suggests that its autopolysialylation could be involved in promoting favorable NCAM interactions or in modulating coordination or scaffolding of the sugar acceptor. This may explain the observation that NCAM Ig5 muta tions of Ser448 and Asn476 (immediately adjacent to NCAM poly sialylation sites Asn449 and Asn478) to the equivalently positioned bulky basic residues of the olfactory cell adhesion molecule abrogates polysialylation 55 , potentially by affecting the interaction with the Asn74linked autopolysialylated glycan. As docked, NCAM Ig5 Asn449 and Asn478 are slightly farther from the ST8SiaIV active site than Asn74 (~50-55 Å, Fig.…”

Section: A Model Of Polysialyltransferase-acceptor Interactionsmentioning

confidence: 95%

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Structure of human ST8SiaIII sialyltransferase provides insight into cell-surface polysialylation

Volkers

Worrall

Kwan

et al. 2015

Nat Struct Mol Biol

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Sialyltransferases of the mammalian ST8Sia family catalyze oligo- and polysialylation of surface-localized glycoproteins and glycolipids through transfer of sialic acids from CMP-sialic acid to the nonreducing ends of sialic acid acceptors. The crystal structure of human ST8SiaIII at 1.85-Å resolution presented here is, to our knowledge, the first solved structure of a polysialyltransferase from any species, and it reveals a cluster of polysialyltransferase-specific structural motifs that collectively provide an extended electropositive surface groove for binding of oligo-polysialic acid chain products. The ternary complex of ST8SiaIII with a donor sugar analog and a sulfated glycan acceptor identified with a sialyltransferase glycan array provides insight into the residues involved in substrate binding, specificity and sialyl transfer.

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Section: A Model Of Polysialyltransferase-acceptor Interactionsmentioning

confidence: 54%

Section: A Model Of Polysialyltransferase-acceptor Interactionsmentioning

confidence: 95%

Structure of human ST8SiaIII sialyltransferase provides insight into cell-surface polysialylation

Volkers

Worrall

Kwan

et al. 2015

Nat Struct Mol Biol

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“…While we cannot rule out this possibility, analyses of NCAM chimeras and mutants highlighted the importance of the composition and length of the Ig5–FN1 linker region and secondary interactions between the polyST docked on the FN1 acidic patch and the Ig5 domain. 29,48 …”

Section: Discussionmentioning

confidence: 99%

“…The OCAM FN1 domain could be recognized by ST8Sia-IV, but large basic residues in the OCAM Ig5 domain blocked polyST access and N -glycan polysialylation, suggesting that the polySTs make contact with amino acid sequences in this NCAM domain that are required for their proper positioning. 29 Consequently, changes in the Ig5–FN1 linker or the GGVPI or NGKG loop regions that stabilize the linker or even sequences adjacent to the linker, like PSSP, may alter the Ig5–FN1 domain orientation and block this secondary binding interaction in the Ig5 domain.…”

Section: Discussionmentioning

confidence: 99%

The Polybasic Region of the Polysialyltransferase ST8Sia-IV Binds Directly to the Neural Cell Adhesion Molecule, NCAM

et al. 2017

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Polysialic acid (polySia) is a unique post-translational modification found on a small set of mammalian glycoproteins. Composed of long chains of α2,8-linked sialic acid, this large, negatively charged polymer attenuates protein and cell adhesion and modulates signaling mediated by its carriers and proteins that interact with these carriers. PolySia is crucial for the proper development of the nervous system and is upregulated during tissue regeneration and in highly invasive cancers. Our laboratory has previously shown that the neural cell adhesion molecule, NCAM, has an acidic surface patch in its first fibronectin type III repeat (FN1) that is critical for the polysialylation of N-glycans on the adjacent immunoglobulin domain (Ig5). We have also identified a polysialyltransferase (polyST) polybasic region (PBR) that may mediate substrate recognition. However, a direct interaction between the NCAM FN1 acidic patch and the polyST PBR has yet to be demonstrated. Here, we have probed this interaction using isothermal titration calorimetry and nuclear magnetic resonance (NMR) spectroscopy. We observe direct and specific binding between FN1 and the PBR peptide that is dependent upon acidic residues in FN1 and basic residues of the PBR. NMR titration experiments verified the role of the FN1 acidic patch in the recognition of the PBR and suggest a conformational change of the Ig5–FN1 linker region following binding of the PBR to the acidic patch. Finally, mutation of residues identified by NMR titration experiments impacts NCAM polysialylation, supporting their mechanistic role in protein-specific polysialylation.

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“…Evidence clearly demonstrates that polysialylation is a protein-specific reaction in which the polySTs initially recognise sequences on NCAM prior to effecting glycan modifications [14–17]. The polySTs show independent but overlapping expression patterns [18–20], with ST8SiaII being the prominent enzyme in embryonic development [19,21].…”

Section: Introductionmentioning

confidence: 99%

Pharmacological Inhibition of polysialyltransferase ST8SiaII Modulates Tumour Cell Migration

et al. 2013

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Polysialic acid (polySia), an α-2,8-glycosidically linked polymer of sialic acid, is a developmentally regulated post-translational modification predominantly found on NCAM (neuronal cell adhesion molecule). Whilst high levels are expressed during development, peripheral adult organs do not express polySia-NCAM. However, tumours of neural crest-origin re-express polySia-NCAM: its occurrence correlates with aggressive and invasive disease and poor clinical prognosis in different cancer types, notably including small cell lung cancer (SCLC), pancreatic cancer and neuroblastoma. In neuronal development, polySia-NCAM biosynthesis is catalysed by two polysialyltransferases, ST8SiaII and ST8SiaIV, but it is ST8SiaII that is the prominent enzyme in tumours. The aim of this study was to determine the effect of ST8SiaII inhibition by a small molecule on tumour cell migration, utilising cytidine monophosphate (CMP) as a tool compound. Using immunoblotting we showed that CMP reduced ST8iaII-mediated polysialylation of NCAM. Utilizing a novel HPLC-based assay to quantify polysialylation of a fluorescent acceptor (DMB-DP3), we demonstrated that CMP is a competitive inhibitor of ST8SiaII (K i = 10 µM). Importantly, we have shown that CMP causes a concentration-dependent reduction in tumour cell-surface polySia expression, with an absence of toxicity. When ST8SiaII-expressing tumour cells (SH-SY5Y and C6-STX) were evaluated in 2D cell migration assays, ST8SiaII inhibition led to significant reductions in migration, while CMP had no effect on cells not expressing ST8SiaII (DLD-1 and C6-WT). The study demonstrates for the first time that a polysialyltransferase inhibitor can modulate migration in ST8SiaII-expressing tumour cells. We conclude that ST8SiaII can be considered a druggable target with the potential for interfering with a critical mechanism in tumour cell dissemination in metastatic cancers.

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The Polysialyltransferases Interact with Sequences in Two Domains of the Neural Cell Adhesion Molecule to Allow Its Polysialylation

Cited by 16 publications

References 44 publications

Structure of human ST8SiaIII sialyltransferase provides insight into cell-surface polysialylation

Structure of human ST8SiaIII sialyltransferase provides insight into cell-surface polysialylation

The Polybasic Region of the Polysialyltransferase ST8Sia-IV Binds Directly to the Neural Cell Adhesion Molecule, NCAM

Pharmacological Inhibition of polysialyltransferase ST8SiaII Modulates Tumour Cell Migration

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