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Dimitrios Krikorian

Publications and source records attributed to Dimitrios Krikorian.

8 recordsLinked to original sources
PubMed2006

Phosphorylated and nonphosphorylated epitopes of the La/SSB autoantigen: comparison of their antigenic and conformational characteristics.

La/SSB phosphoprotein is the target antigen of autoantibodies in sera of patients with Sjögren's syndrome (SS) and systemic lupus erythematosus (SLE). Among other structural and function motifs, four phosphorylation sites are encompassed in the primary sequence of La/SSB. Two of them (Thr-362 and Ser-366) are located within GSGKGKVQFQGKKTKFASDD (346-368) and one (Thr-302) within VTWEVLEGEVEKEALKKI (301-318), which are main B-cell epitopes of La/SSB. With the aim to investigate how phosphorylation, one of the most common posttranslational protein modifications, affects the antigenic and conformational characteristics of the La/SSB epitopes, we synthesized and studied the phosphorylated epitopes La/SSB(346-368)-P, La/SSB(359-368)-P, and La/SSB(301-318)-P with respect to their nonphosphorylated counterparts. Anti-La/SSB positive sera from SS and SLE patients are better recognized by the phosphorylated epitopes compared to their nonphosphorylated counterparts. Conformational analysis by (1)H nuclear magnetic resonance spectroscopy and molecular dynamics showed that the phosphorylated epitopes adopt different structural characteristics from those of the corresponding nonphosphorylated epitopes. It is concluded that phosphorylation can create neoepitopes with altered functions, compared to the nonphosphorylated epitopes, which might be seen from the immune system as "foreign."

Amino Acid Sequence↗
PubMed2006

Influence of sequential oligopeptide carriers on the bioactive structure of conjugated epitopes: comparative study of the conformation of a Herpes simplex virus glycoprotein gD-1 epitope in the free and conjugated form, and protein "built-in" crystal structure.

Synthetic carriers play an important role in immunogen presentation, due to their ability of inducing improved and specific responses to conjugated epitopes. Their influence on the bioactive conformation of the epitope, though admittedly crucial for relevant in vitro and in vivo applications, is difficult to evaluate, given the usual lack of information on the complex conformational features determined by the nature of the carrier and the mode of ligation. Using the Herpes simplex virus glycoprotein D-1 epitope (Leu(9)-Lys-Nle-Ala-Asp-Pro-Asn-Arg-Phe-Arg-Gly-Lys-Asp-Leu(22)) as a model, we have performed a detailed conformational analysis on the free epitope peptide in solution and on three constructs in which the epitope was conjugated to sequential oligopeptide carriers {Ac-[Lys-Aib-Gly](4)-OH (SOC(4))} (through either a thioether or an amide bond; Ac: acetyl) and polytuftsin oligomers {H-[Thr-Lys-Pro-Lys-Gly](4)-NH(2) (T20)}, (through a thioether bond). The analysis of the epitope conformation in the parent protein, in carrier-conjugated and free form, suggests that the beta-turn structure of the -Asp(13)-Pro-Asn-Arg(16)- segment is highly conserved and independent of the epitope form. However, small conformational variations were observed at the C-terminal part of the epitope, depending on the nature of the carrier.

Amino Acid Sequence↗
PubMed2006

Artificial carriers: a strategy for constructing antigenic/immunogenic conjugates.

The rational design of artificial carriers for anchoring multiple copies of B and/or T cell epitopes, built-in vaccine adjuvants and "promiscuous" T cell epitopes for the construction of conjugates as antigenic substrates or potent immunogens has been the stimulus of intensive efforts nowadays. The unambiguous composition, the reliability and the versatility of the production of reconstituted antigens or immunogens has found a great number of biochemical applications in developing immunoassays of high sensitivity, specificity and reproducibility and in generating site-specific antibodies for usage as human vaccine candidates. In this review are summarized different types of artificial carriers currently used as dendrimers bearing branching segments, multimeric core matrices and templates with built-in folding devices. Emphasis is given to the construction and application of a helicoid-type Sequential Oligopeptide Carrier (SOCn) developed in our laboratory. The beneficial structural elements of SOCn induce a favorable arrangement of the conjugated peptides, which also retain their initial "active" conformation, so that potent antigens and immunogens are generated.

Animals↗
PubMed2005

Innovative, multifunctional sequential oligopeptide carriers Soc(n)-I and SOC(n)-II: functions-technology-perspectives.

An innovative type of multifunctional helicoid artificial carriers, formed by the repetitive Lys-Aib-Gly (SOC(n)-I) or by the Aib-Lys-Aib-Gly (SOC(n)-II), with structural rigidity and regularity were successfully applied in our laboratory for anchoring antigenic/immunogenic peptides. The carriers, designed to display a predetermined 3D structure, adopt the 3(10) helical conformation, while the attached peptides preserve their original "active" conformation. The constructed conjugates were used as substrates in solid phase immunoassays, as well as for generating potent and specific immune responses.

Amino Acid Sequence↗
PubMed2003

Solution conformation of the antibody-bound tyrosine phosphorylation site of the nicotinic acetylcholine receptor beta-subunit in its phosphorylated and nonphosphorylated states.

Phosphorylation of the acetylcholine receptor (AChR) seems to be responsible for triggering several effects including its desensitization and aggregation at the postsynaptic membrane and probably initiates a signal transduction pathway at the postsynaptic membrane. To study the structural and functional role of the tyrosine phosphorylation site of the AChR beta-subunit and contribute to the in-depth understanding of the structural basis of the ion channel function, we synthesized four peptides containing the phosphorylated and nonphosphorylated sequences (380-391) of the human and Torpedo AChR beta-subunits and studied their interaction with a monoclonal antibody (mAb 148) that is known to bind to this region and that is capable of blocking ion channel function. All four peptides were efficient inhibitors of mAb 148 binding to AChR, although the nonphosphorylated human peptide was considerably less effective than the three others. We then investigated the conformation acquired by all four peptides in their antibody-bound state, which possibly illustrates the local conformation of the corresponding sites on the intact AChR molecule. The phosphorylated human and Torpedo peptides adopted a distorted 3(10) helix conformation. The nonphosphorylated Torpedo peptide, which is also an efficient inhibitor, was also folded. In contrast, the nonphosphorylated human peptide (a less efficient inhibitor) presented an extended structure. It is concluded that the phosphorylation of the AChR at its beta-subunit Tyr site leads to a significant change in its conformation, which may affect several functions of the AChR.

Amino Acid Sequence↗
PubMedDate not supplied

Synthesis and comparison of antibody recognition of conjugates containing herpes simplex virus type 1 glycoprotein D epitope VII.

Synthetic oligopeptides comprising linear or continuous topographic B-cell epitope sequences of proteins might be considered as specific and small size antigens. It has been demonstrated that the strength and specificity of antibody binding could be altered by conjugation to macromolecules or by modification in the flanking regions. However, no systematic studies have been reported to describe the effect of different carrier macromolecules in epitope conjugates. To this end, the influence of carrier structure and topology on antibody recognition of attached epitope has been studied by comparing the antibody binding properties of a new set of conjugates with tetratuftsin analogue (H-[Thr-Lys-Pro-Lys-Gly](4)-NH(2), T20) sequential oligopeptide carrier (SOC(n)), branched chain polypeptide, poly[Lys(Ser(i)-DL-Ala(m))] (SAK), multiple antigenic peptide (MAP), and keyhole limpet hemocyanine (KLH). In these novel constructs, peptide (9)LKNleADPNRFRGKDL(22) ([Nle(11)]-9-22) representing an immunodominant B cell epitope of herpes simplex virus type 1 glycoprotein D (HSV-1 gD) was conjugated to polypeptides through a thioether or amide bond. Here we report on the preparation of sequential and polymeric polypeptides possessing chloroacetyl groups in multiple copies at the alpha- and/or epsilon-amino group of the polypeptides and its use for the conjugation of epitope peptides possessing Cys at C-terminal position. We have performed binding studies (direct and competitive ELISA) with monoclonal antibody (Mab) A16, recognizing the HSV gD-related epitope, [Nle(11)]-9-22, and conjugates containing identical and uniformly oriented epitope peptide in multiple copies attached to five different macromolecules as carrier. Data suggest that the chemical nature of the carrier and the degree of substitution have marked influence on the strength of antibody binding.

Amino Acid Sequence↗
PubMedDate not supplied

A peptide carrier with a built-in vaccine adjuvant: construction of immunogenic conjugates.

A multifunctional carrier combining B/T cell epitopes (i), a built-in vaccine adjuvant (ii), and a universal T cell epitope (iii) for the construction of potent and specific immunogenic conjugates is presented. The IL-1beta(163-171) fragment known to reproduce the immunostimulatory and adjuvant effects of the whole IL-1beta without possessing any of the pro-inflammatory properties of IL-1beta was covalently anchored to the N-terminus of the Sequential Oligopeptide Carrier, SOC(n), formed by the repeating tripeptide unit Lys-Aib-Gly. A promiscuous T cell epitope derived from the tetanus toxin, TT(593-599), was also positioned in the carboxy terminus of SOC(n) as a universal immunogen to provide broad immunogenicity. Selected B/T cell epitopes from the Sm and La/SSB autoantigens, against which is directed the humoral autoimmunity in patients with systemic lupus erythematosus and Sjögren's Syndrome, respectively, were coupled to the Lys-N(epsilon)H2 groups of the carrier, and the formulated constructs were administered in animals following the conventional immunization protocol of complete/incomplete Freund's adjuvant. The induced immune responses were compared with that produced when the Sm- and La/SSB-reconstituted immunogenic conjugates were injected alone. High titers of specific antibodies recognizing the priming construct, as well as the cognate autoantigen, were obtained when administered alone without the assistance of Freund's adjuvant. It is concluded that our approach provides the conceptual and experimental framework for the development of multifunctional immunogenic conjugates eliciting enhanced, specific, and prolonged humoral response for usage as human vaccine candidates.

Adjuvants, Immunologic↗