Chemoenzymatic Synthesis of HIV‐1 gp41 Glycopeptides: Effects of Glycosylation on the Anti‐HIV Activity and α‐Helix Bundle‐Forming Ability of Peptide C34

Wang, Lai-Xi; Song, Haijing; Liu, Shuwen; Liang, Hong; Jiang, Shibo; Ni, Jiahong; Li, Hengguang

doi:10.1002/cbic.200400440

Cited by 62 publications

(55 citation statements)

References 49 publications

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“…Several studies also described an improved thermal stability and/or improved free energy of the 6-HB with the secondary substitutions in the CHR region (8,53,58,59). Interestingly, the removal of the glycan at position 126 can also enhance the thermal stability of the NHR-CHR complex (60). In this study, we found again that a single N126K substitution could render the virus mildly resistant to diverse inhibitors (ϳ3-fold), which emphasized that the mechanism of N126K-mediated resistance is an enhanced viral 6-HB rather than direct binding.…”

Section: Figmentioning

confidence: 99%

Mechanism of HIV-1 Resistance to Short-Peptide Fusion Inhibitors Targeting the Gp41 Pocket

Sai

Chong

Qiu

et al. 2015

J Virol

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The deep hydrophobic pocket on the N trimer of HIV-1 gp41 has been considered an ideal drug target. On the basis of the M-T hook structure, we recently developed short-peptide-based HIV-1 fusion inhibitors (MTSC22 and HP23), which mainly target the pocket site and possess highly potent antiviral activity. In this study, we focused on investigating their resistance pathways and mechanisms by escape HIV-1 mutants to SC22EK, a template peptide for MTSC22 and HP23. Two substitutions, E49K and N126K, located, respectively, at the N-and C-heptad repeat regions of gp41, were identified as conferring high resistance to the inhibitors targeting the pocket and cross-resistance to enfuvirtide (T20) and sifuvirtide (SFT). The underlying mechanisms of SC22EK-induced resistance include the following: (i) significantly reduced binding affinity of the inhibitors, (ii) dramatically enhanced interaction of the viral six-helix bundle, and (iii)severely damaged functionality of the viral Env complex. Our data have provided important information for the structure-function relationship of gp41 and the structure-activity relationship of viral fusion inhibitors. IMPORTANCEEnfuvirtide (T20) is the only HIV-1 fusion inhibitor in clinical use, but the problem of resistance significantly limits its use, calling for new strategies or concepts to develop next-generation drugs. On the basis of the M-T hook structure, short-peptide HIV-1 fusion inhibitors specifically targeting the gp41 pocket site exhibit high binding and antiviral activities. Here, we investigated the molecular pathway of HIV-1 resistance to the short inhibitors by selecting and mapping the escape mutants. The key substitutions for resistance and the underlying mechanisms have been finely characterized. The data provide important information for the structure-function relationship of gp41 and its inhibitors and will definitely help our future development of novel drugs that block gp41-dependent fusion.

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Section: Figmentioning

confidence: 99%

Mechanism of HIV-1 Resistance to Short-Peptide Fusion Inhibitors Targeting the Gp41 Pocket

Sai

Chong

Qiu

et al. 2015

J Virol

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“…We have applied the N175A mutant for the synthesis of Man 9 GlcNAc 2 -C34, a 34-mer glycopeptide derived from HIV-1 gp41 that carries a native high mannose N-glycan (Scheme 4). Man 9 GlcNAc 2 -C34 was shown to be a potent HIV entry inhibitor (IC 90 ϭ 12 nM) (35). In contrast to the parent polypeptide C34, Man 9 GlcNAc 2 -C34 has much higher water solubility than C34 (35), thus having a potential to be developed as an anti-HIV drug.…”

Section: Site-directed Mutagenesis Of the Residues In The Putative Camentioning

confidence: 99%

“…Man 9 GlcNAc 2 -C34 was shown to be a potent HIV entry inhibitor (IC 90 ϭ 12 nM) (35). In contrast to the parent polypeptide C34, Man 9 GlcNAc 2 -C34 has much higher water solubility than C34 (35), thus having a potential to be developed as an anti-HIV drug. The glycopeptide was previously synthesized in about 10% yield by Endo-A-catalyzed transglycosylation using 5-fold excess of Man 9 GlcNAc 2 -Asn as the donor substrate (35).…”

Section: Site-directed Mutagenesis Of the Residues In The Putative Camentioning

confidence: 99%

“…In contrast to the parent polypeptide C34, Man 9 GlcNAc 2 -C34 has much higher water solubility than C34 (35), thus having a potential to be developed as an anti-HIV drug. The glycopeptide was previously synthesized in about 10% yield by Endo-A-catalyzed transglycosylation using 5-fold excess of Man 9 GlcNAc 2 -Asn as the donor substrate (35). Here we found that when Man 9 GlcNAc-oxazoline was used as the donor substrate, the N175A-catalyzed transglycosylation to GlcNAc-C34 (9) proceeded smoothly to give a much higher yield of the product Man 9 GlcNAc 2 -C34 (10) (72%), using only a 2-fold excess of the donor substrate.…”

Section: Site-directed Mutagenesis Of the Residues In The Putative Camentioning

confidence: 99%

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Mutants of Mucor hiemalis Endo-β-N-acetylglucosaminidase Show Enhanced Transglycosylation and Glycosynthase-like Activities

Umekawa

Huang

et al. 2008

Journal of Biological Chemistry

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216

229

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Endo-␤-N-acetylglucosaminidase from Mucor hiemalis (Endo-M), a family 85 glycoside hydrolase, acts on the ␤1,4 linkage of N,N-diacetylchitobiose moiety in the N-linked glycans of glycoproteins and catalyzes not only the hydrolysis reaction but also the transglycosylation reaction that transfers the releasing sugar chain to an acceptor other than water to form a new glycosidic linkage. The transglycosylation activity of Endo-M holds a great promise for the chemo-enzymatic synthesis and glycoengineering of glycoproteins, but the inherent hydrolytic activity for product hydrolysis and low transglycosylation have hampered its broad applications. This paper describes the sitedirected mutagenesis on residues in the putative catalytic region of Endo-M to generate mutants with superior transglycosylation activity. Two interesting mutants were discovered. The Y217F mutant was found to possess much enhanced transglycosylation activity and yet much diminished hydrolytic activity in comparison with the wild-type Endo-M. Kinetic analyses revealed that the K m value of Y217F for an acceptor substrate 4-methylumbelliferyl-␤-D-N-acetylglucosaminide was only one-tenth of that of the wild-type, implicating a much higher affinity of Y217F for the acceptor substrate than the wild-type. The other mutant, N175A, acts like a glycosynthase. It was found that mutation at Asn 175 "knocked out" the hydrolytic activity, but the mutant was able to take the highly active sugar oxazolines (the transition state mimics) as donor substrates for transglycosylation. This is the first glycosynthase derived from endo-␤-N-acetylglucosaminidases that proceed via a substrate-assisted mechanism. Our findings provide further insights on the substrate-assisted mechanism of GH85. The usefulness of the novel glycosynthase was exemplified by the efficient synthesis of a human immunodeficiency virus, type 1 (HIV-1) glycopeptide with potent anti-HIV activity.Endo-␤-N-acetylglucosaminidase (EC 3.2.1.96) (ENGase) 3 catalyzes hydrolysis of the ␤1,4-glycosidic linkage of the N,NЈ-diacetylchitobiose moiety in the core of asparagine-linked glycan of various glycoproteins and glycopeptides. This type of enzyme is widely distributed in animals, plants, fungi, and bacteria. Several bacterial enzymes, such as Endo-H from Streptomyces plicatus (1) and Endo-F 1 from Flavobacterium meningosepticum (2), were cloned and classified into glycoside hydrolase (GH) family 18 in the CAZy data base (available on the World Wide Web), which may share a common evolutional origin with GH18 chitinases. The other ENGases are distinct from the enzymes of the GH18 chitinase family and are classified into the GH family 85. We and others have previously reported that several ENGases of the GH85 family showed significant transglycosylation activity (i.e. the ability to transfer the releasing glycan to an acceptor other than water to form a new glycosidic linkage) (3-6). These ENGases include Endo-M from Mucor hiemalis (3), Endo-A from Arthrobactor protophormiae (4), Endo-CE from Caenorhabditi...

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“…It has been demonstrated that oligomannose modification of the asparagine residue of C34, a 34-mer peptide derived from the C-ectodomain of HIV-1 envelope glycoprotein, dramatically improved its solubility and stability as a promising candidate for anti-HIV agents (65,66). Based on the hydrophilic outside surface and the hydrophobic internal cavity, cyclodextrins have been successfully exploited to form inclusion complexes with various hydrophobic drugs to dramatically improve their water solubility (67).…”

Section: Carbohydrates Mediated Drug Modification and Deliverymentioning

confidence: 99%

Carbohydrate drugs: current status and development prospect

Zhang

Wang

2015

DD&T

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Chemoenzymatic Synthesis of HIV‐1 gp41 Glycopeptides: Effects of Glycosylation on the Anti‐HIV Activity and α‐Helix Bundle‐Forming Ability of Peptide C34

Cited by 62 publications

References 49 publications

Mechanism of HIV-1 Resistance to Short-Peptide Fusion Inhibitors Targeting the Gp41 Pocket

Mechanism of HIV-1 Resistance to Short-Peptide Fusion Inhibitors Targeting the Gp41 Pocket

Mutants of Mucor hiemalis Endo-β-N-acetylglucosaminidase Show Enhanced Transglycosylation and Glycosynthase-like Activities

Carbohydrate drugs: current status and development prospect

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