Solid-phase synthesis and applications of N-(S-acetylmercaptoacetyl) peptides

Drijfhout, Jan W.; Bloemhoff, W.; Poolman, Jan; Hoogerhout, P.

doi:10.1016/0003-2697(90)90468-o

Cited by 21 publications

(7 citation statements)

References 11 publications

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“…Total IgG concentration in each serum sample was adjusted to 9.3 mg IgG/mL and serum specific IgG titers against WH7 or LTA were measured by ELISA. Nunc-immuno Maxisorp MicroWell 96 well plates were coated either with 0.2 µg of WH7 antigen (previously conjugated with Tetanus Toxoid as described by Hoogerhout and co-workers [25] , [26] ) or 0.125 µg of LTA from S. aureus purchased from Sigma (St. Louis, Mo.) in 0.2M carbonate-bicarbonate coating buffer.…”

Section: Methodsmentioning

confidence: 99%

Synthetic Teichoic Acid Conjugate Vaccine against Nosocomial Gram-Positive Bacteria

et al. 2014

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Lipoteichoic acids (LTA) are amphiphilic polymers that are important constituents of the cell wall of many Gram-positive bacteria. The chemical structures of LTA vary among organisms, albeit in the majority of Gram-positive bacteria the LTAs feature a common poly-1,3-(glycerolphosphate) backbone. Previously, the specificity of opsonic antibodies for this backbone present in some Gram-positive bacteria has been demonstrated, suggesting that this minimal structure may be sufficient for vaccine development. In the present work, we studied a well-defined synthetic LTA-fragment, which is able to inhibit opsonic killing of polyclonal rabbit sera raised against native LTA from Enterococcus faecalis 12030. This promising compound was conjugated with BSA and used to raise rabbit polyclonal antibodies. Subsequently, the opsonic activity of this serum was tested in an opsonophagocytic assay and specificity was confirmed by an opsonophagocytic inhibition assay. The conjugated LTA-fragment was able to induce specific opsonic antibodies that mediate killing of the clinical strains E. faecalis 12030, Enterococcus faecium E1162, and community-acquired Staphylococcus aureus strain MW2 (USA400). Prophylactic immunization with the teichoic acid conjugate and with the rabbit serum raised against this compound was evaluated in active and passive immunization studies in mice, and in an enterococcal endocarditis rat model. In all animal models, a statistically significant reduction of colony counts was observed indicating that the novel synthetic LTA-fragment conjugate is a promising vaccine candidate for active or passive immunotherapy against E. faecalis and other Gram-positive bacteria.

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Section: Methodsmentioning

confidence: 99%

Synthetic Teichoic Acid Conjugate Vaccine against Nosocomial Gram-Positive Bacteria

et al. 2014

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“…Polyclonal antiserum #32 against amino acids 61 to 77 of EBV gp42 was raised in a rabbit by using synthetic peptide VNFNKTA EQEYGDKEVK conjugated to tetanus toxoid (6).…”

Section: Methodsmentioning

confidence: 99%

Epstein-Barr Virus gp42 Is Posttranslationally Modified To Produce Soluble gp42 That Mediates HLA Class II Immune Evasion

Ressing¹,

Leeuwen²,

Verreck

et al. 2005

J Virol

100

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Epstein-Barr virus (EBV) resides as a persistent infection in human leukocyte antigen (HLA) class II؉ B lymphocytes and is associated with a number of malignancies. The EBV lytic-phase protein gp42 serves at least two functions: gp42 acts as the coreceptor for viral entry into B cells and hampers T-cell recognition via HLA class II molecules through steric hindrance of T-cell receptor-class II-peptide interactions. Here, we show that gp42 associates with class II molecules at their various stages of maturation, including immature ␣␤Ii heterotrimers and mature ␣␤-peptide complexes. When analyzing the biosynthesis and maturation of gp42 in cells stably expressing the viral protein, we found that gp42 occurs in two forms: a full-length type II membrane protein and a truncated soluble form. Soluble gp42 is generated by proteolytic cleavage in the endoplasmic reticulum and is secreted. Soluble gp42 is sufficient to inhibit HLA class II-restricted antigen presentation to T cells. In an almost pure population of Burkitt's lymphoma cells in the EBV lytic cycle, both transmembrane and soluble forms of gp42 are detected. These results imply that soluble gp42 is generated during EBV lytic infection and could contribute to undetected virus production by mediating evasion from T-cell immunity. Epstein-Barr virus (EBV)is a large DNA virus belonging to the family Herpesviridae, subfamily Gammaherpesviriniae, genus Lymphocryptovirus. Like other herpesviruses, EBV persists for life, establishing a latent infection in B lymphocytes with occasional viral reactivation (29). Approximately 90% of the adult population carries EBV DNA. EBV is the causative agent of infectious mononucleosis and is associated with malignancies that originate from lymphoid cells (e.g., Burkitt's lymphoma and Hodgkin's lymphoma) and epithelial cells (e.g., nasopharyngeal carcinoma). In immunosuppressed or immunocompromised individuals, EBV can cause (fatal) lymphoproliferative disease. In contrast, in healthy individuals, EBV is well controlled by the immune system. The widespread and mostly asymptomatic persistent EBV infections in adults reflect the balance between viral replication and host immune control.EBV dedicates part of its genome to immune evasion functions (15). Modulation of T-cell recognition is an important target for EBV, as the virus resides intracellularly for most of its life cycle. Fragments of viral proteins can be displayed at the cell surface by human leukocyte antigen (HLA) class I and class II molecules for the activation of T cells carrying receptors of the appropriate specificity. Activated T cells may then lead to elimination of the pathogen, among other ways through destruction of the infeced cell. To escape from antiviral immunity, EBV should interfere with both CD8 ϩ and CD4 ϩ T-cell responses, particularly as the virus infects B lymphocytes expressing both classes of HLA molecules. The EBV nuclear protein EBNA1 contains a glycine-alanine repeat domain that renders the protein resistant to proteasomal degradation and inhibits...

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“…However, if one wishes to synthesize a peptide in order to prepare a conjugate, planning the synthesis for this purpose can prove very advantageous. For example, some workers have introduced a reactive cysteine residue at the desired position for heteroligation (11,12); Drijfhout et al (13) completed a sequence on a solid support with an N-terminal S-acetylmercaptoacetyl group. The deprotected peptide was treated with hydroxylamine to remove the S-acetyl group and then joined through its N-terminal sulfhydryl group to a conjugate partner bearing an SH-selective electrophilic function (e.g., an N-substituted maleimide or an a-haloacetyl moiety) by means of a very stable thioether linkage (12,13).…”

Section: Introductionmentioning

confidence: 99%

Synthesis of N.alpha.-(tert-butoxycarbonyl)-N.epsilon.-[N-(bromoacetyl)-.beta.-alanyl]-L-lysine: Its use in peptide synthesis for placing a bromoacetyl cross-linking function at any desired sequence position

Inman

Highet²,

Kolodny³

et al. 1991

Bioconjugate Chem.

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A new amino acid derivative, N alpha-(tert-butoxycarbonyl)-N epsilon-[N-(bromoacetyl)-beta-alanyl]-L-lysine (BBAL), has been synthesized as a reagent to be used in solid-phase peptide synthesis for introducing a side-chain bromoacetyl group at any desired position in a peptide sequence. The bromoacetyl group subsequently serves as a sulfhydryl-selective cross-linking function for the preparation of cyclic peptides, peptide conjugates, and polymers. BBAL is synthesized by condensation of N-bromoacetyl-beta-alanine with N alpha-Boc-L-lysine and is a white powder which is readily stored, weighed, and used with a peptide synthesizer, programmed for N alpha-Boc amino acid derivatives. BBAL residues are stable to final HF deprotection/cleavage. BBAL peptides can be directly coupled to other molecules or surfaces which possess free sulfhydryl groups by forming stable thioether linkages. Peptides containing both BBAL and cysteine residues can be self-coupled to produce either cyclic molecules or linear peptide polymers, also linked through thioether bonds. Products made with BBAL peptides may be characterized by amino acid analysis of acid hydrolyzates by quantification of beta-alanine, which separates from natural amino acids in suitable analytical systems. Where sulfhydryl groups on coupling partners arise from cysteine residues, S-(carboxymethyl)cysteine in acid hydrolyzates may also be assayed for this purpose. Examples are given of the use of BBAL in preparing peptide polymers and a peptide conjugate with bovine albumin to serve as immunogens or model vaccine components.

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Solid-phase synthesis and applications of N-(S-acetylmercaptoacetyl) peptides

Cited by 21 publications

References 11 publications

Synthetic Teichoic Acid Conjugate Vaccine against Nosocomial Gram-Positive Bacteria

Synthetic Teichoic Acid Conjugate Vaccine against Nosocomial Gram-Positive Bacteria

Epstein-Barr Virus gp42 Is Posttranslationally Modified To Produce Soluble gp42 That Mediates HLA Class II Immune Evasion

Synthesis of N.alpha.-(tert-butoxycarbonyl)-N.epsilon.-[N-(bromoacetyl)-.beta.-alanyl]-L-lysine: Its use in peptide synthesis for placing a bromoacetyl cross-linking function at any desired sequence position

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