Structures of activin ligand traps using natural sets of type I and type II TGFß receptors.

The 30+ unique ligands of the TGFß family signal by forming complexes using different combinations of type I and type II receptors. Therapeutically, the extracellular domain of a single receptor fused to an Fc molecule can effectively neutralize subsets of ligands. Increased ligand specificity can be accomplished by using the extracellular domains of both the type I and type II receptor to mimic the naturally occurring signaling complex. Here, we report the structure of one "type II-type I-Fc" fusion, ActRIIB-Alk4-Fc, in complex with two TGFß family ligands, ActA, and GDF11, providing a snapshot of this therapeutic platform. The study reveals that extensive contacts are formed by both receptors, replicating the ternary signaling complex, despite the inherent low affinity of Alk4. Our study shows that low-affinity type I interactions support altered ligand specificity and can be visualized at the molecular level using this platform.

Atypical haemolytic uremic syndrome with posterior reversible encephalopathy syndrome in an adolescent: a rare case report.

Atypical hemolytic uremic syndrome (aHUS) is a group of disorders that affect kidneys which is rare type of HUS that differs from classical hemolytic uremic syndrome (HUS) by absence of prodromal phase consisting of episodes of diarrhoea due to preceding shiga toxin E. coli (STEC-HUS) infection and is 5% of all HUS cases. Approximately 50% cases present with clinical triad of hemolytic anemia, thrombocytopenia and renal insufficiency. However, it can have unusual clinical features in form of central nervous system involvement. This case, of a 15-year-old Indian boy, is one such rare presentation of atypical haemolytic uremic syndrome associated with posterior reversible encephalopathy syndrome (PRES), or reversible posterior leukoencephalopathy syndrome (RPLS) who presented with anaemia, anasarca, papilledema, hypertension, episodic seizures and significant magnetic resonance imaging (MRI) brain findings. We report this uncommon combination of two syndromes to provide useful insight for clinicians to approach and diagnose such presentation in paediatric patients.

Functionally diverse heteromeric traps for ligands of the transforming growth factor-ß superfamily.

Ligands of the transforming growth factor-ß (TGF-ß) superfamily are important targets for therapeutic intervention but present challenges because they signal combinatorially and exhibit overlapping activities in vivo. To obtain agents capable of sequestering multiple TGF-ß superfamily ligands with novel selectivity, we generated soluble, heterodimeric ligand traps by pairing the extracellular domain (ECD) of the native activin receptor type IIB (ActRIIB) alternately with the ECDs of native type I receptors activin receptor-like kinase 4 (ALK4), ALK7, or ALK3. Systematic analysis of these heterodimeric constructs by surface plasmon resonance, and comparison with their homodimeric counterparts, revealed that each type I receptor partner confers a distinct ligand-binding profile to the heterodimeric construct. Additional characterization in cell-based reporter gene assays confirmed that the heterodimeric constructs possessed different profiles of signaling inhibition in vitro, which translated into altered patterns of pharmacological activity when constructs were administered systemically to wild-type mice. Our results detail a versatile platform for the modular recombination of naturally occurring receptor domains, giving rise to inhibitory ligand traps that could aid in defining the physiological roles of TGF-ß ligand sets or be directed therapeutically to human diseases arising from dysregulated TGF-ß superfamily signaling.

ActRIIB:ALK4-Fc alleviates muscle dysfunction and comorbidities in murine models of neuromuscular disorders.

Patients with neuromuscular disorders suffer from a lack of treatment options for skeletal muscle weakness and disease comorbidities. Here, we introduce as a potential therapeutic agent a heterodimeric ligand-trapping fusion protein, ActRIIB:ALK4-Fc, which comprises extracellular domains of activin-like kinase 4 (ALK4) and activin receptor type IIB (ActRIIB), a naturally occurring pair of type I and II receptors belonging to the TGF-ß superfamily. By surface plasmon resonance (SPR), ActRIIB:ALK4-Fc exhibited a ligand binding profile distinctly different from that of its homodimeric variant ActRIIB-Fc, sequestering ActRIIB ligands known to inhibit muscle growth but not trapping the vascular regulatory ligand bone morphogenetic protein 9 (BMP9). ActRIIB:ALK4-Fc and ActRIIB-Fc administered to mice exerted differential effects - concordant with SPR results - on vessel outgrowth in a retinal explant assay. ActRIIB:ALK4-Fc induced a systemic increase in muscle mass and function in wild-type mice and in murine models of Duchenne muscular dystrophy (DMD), amyotrophic lateral sclerosis (ALS), and disuse atrophy. Importantly, ActRIIB:ALK4-Fc improved neuromuscular junction abnormalities in murine models of DMD and presymptomatic ALS and alleviated acute muscle fibrosis in a DMD model. Furthermore, in combination therapy ActRIIB:ALK4-Fc increased the efficacy of antisense oligonucleotide M12-PMO on dystrophin expression and skeletal muscle endurance in an aged DMD model. ActRIIB:ALK4-Fc shows promise as a therapeutic agent, alone or in combination with dystrophin rescue therapy, to alleviate muscle weakness and comorbidities of neuromuscular disorders.

Follistatin-288-Fc Fusion Protein Promotes Localized Growth of Skeletal Muscle.

Follistatin is an endogenous glycoprotein that promotes growth and repair of skeletal muscle by sequestering inhibitory ligands of the transforming growth factor-ß superfamily and may therefore have therapeutic potential for neuromuscular diseases. Here, we sought to determine the suitability of a newly engineered follistatin fusion protein (FST288-Fc) to promote localized, rather than systemic, growth of skeletal muscle by capitalizing on the intrinsic heparin-binding ability of the follistatin-288 isoform. As determined by surface plasmon resonance and cell-based assays, FST288-Fc binds to activin A, activin B, myostatin (growth differentiation factor GDF8), and GDF11 with high affinity and neutralizes their activity in vitro. Intramuscular administration of FST288-Fc in mice induced robust, dose-dependent growth of the targeted muscle but not of surrounding or contralateral muscles, in contrast to the systemic effects of a locally administered fusion protein incorporating activin receptor type IIB (ActRIIB-Fc). Furthermore, systemic administration of FST288-Fc in mice did not alter muscle mass or body composition as determined by NMR, which again contrasts with the pronounced systemic activity of ActRIIB-Fc when administered by the same route. Subsequent analysis revealed that FST288-Fc in the circulation undergoes rapid proteolysis, thereby restricting its activity to individual muscles targeted by intramuscular administration. These results indicate that FST288-Fc can produce localized growth of skeletal muscle in a targeted manner with reduced potential for undesirable systemic effects. Thus, FST288-Fc and similar agents may be beneficial in the treatment of disorders with muscle atrophy that is focal, asymmetric, or otherwise heterogeneous.

Integrase residues that determine nucleotide preferences at sites of HIV-1 integration: implications for the mechanism of target DNA binding.

Retroviruses favor target-DNA (tDNA) distortion and particular bases at sites of integration, but the mechanism underlying HIV-1 selectivity is unknown. Crystal structures revealed a network of prototype foamy virus (PFV) integrase residues that distort tDNA: Ala188 and Arg329 interact with tDNA bases, while Arg362 contacts the phosphodiester backbone. HIV-1 integrase residues Ser119, Arg231, and Lys258 were identified here as analogs of PFV integrase residues Ala188, Arg329 and Arg362, respectively. Thirteen integrase mutations were analyzed for effects on integrase activity in vitro and during virus infection, yielding a total of 1610 unique HIV-1 integration sites. Purine (R)/pyrimidine (Y) dinucleotide sequence analysis revealed HIV-1 prefers the tDNA signature (0)RYXRY(4), which accordingly favors overlapping flexible dinucleotides at the center of the integration site. Consistent with roles for Arg231 and Lys258 in sequence specific and non-specific binding, respectively, the R231E mutation altered integration site nucleotide preferences while K258E had no effect. S119A and S119T integrase mutations significantly altered base preferences at positions -3 and 7 from the site of viral DNA joining. The S119A preference moreover mimicked wild-type PFV selectivity at these positions. We conclude that HIV-1 IN residue Ser119 and PFV IN residue Ala188 contact analogous tDNA bases to effect virus integration.

Effectiveness of residual inhibition therapy.

INTRODUCTION: Tinnitus is considered as one of the major symptom associated with many pathologies along with its presence in individuals with normal hearing. With varied cause and mechanism of its generation and increase in number of individuals with complaint of tinnitus, rehabilitation becomes crucial for Audiologists. OBJECTIVE: Study was undertaken to find the efficacy of Residual Inhibition Therapy as treatment procedure for unilateral tinnitus in individuals with normal hearing by comparing pre and post Residual Inhibition Therapy, Contra-lateral Acoustic Reflexes and Tinnitus Handicap Inventory. MATERIALS AND METHODS: Ten subjects between the age range of 20-45 years were included for the study. Tinnitus pitch and intensity were matched and Residual Inhibition Therapy was provided. The Pre Residual Inhibition Therapy, contra-lateral acoustic reflexes and Tinnitus Handicap Inventory scores were compared to post Residual Inhibition Therapy, contra-lateral acoustic reflexes and Tinnitus Handicap Inventory scores. RESULTS AND CONCLUSION: Statistical analysis revealed no significant difference, however, elevated contra-lateral acoustics reflexes post Residual Inhibition Therapy were seen. Even though small sample size made it hard to conclude on the effectiveness of Residual Inhibition Therapy as treatment of tinnitus, elevated contralateral acoustic reflexes post Residual Inhibition Therapy pay way for further advanced studies on the same.

Neural strategies for selective attention distinguish fast-action video game players.

We investigated the psychophysical and neurophysiological differences between fast-action video game players (specifically first person shooter players, FPS) and non-action players (role-playing game players, RPG) in a visual search task. We measured both successful detections (hit rates) and steady-state visually evoked EEG potentials (SSVEPs). Search difficulty was varied along two dimensions: number of adjacent attended and ignored regions (1, 2 and 4), and presentation rate of novel search arrays (3, 8.6 and 20 Hz). Hit rates decreased with increasing presentation rates and number of regions, with the FPS players performing on average better than the RPG players. The largest differences in hit rate, between groups, occurred when four regions were simultaneously attended. We computed signal-to-noise ratio (SNR) of SSVEPs and used partial least squares regression to model hit rates, SNRs and their relationship at 3 Hz and 8.6 Hz. The following are the most significant results: RPG players' parietal responses to the attended 8.6 Hz flicker were predictive of hit rate and were positively correlated with it, indicating attentional signal enhancement. FPS players' parietal responses to the ignored 3 Hz flicker were predictive of hit rate and were positively correlated with it, indicating distractor suppression. Consistent with these parietal responses, RPG players' frontal responses to the attended 8.6 Hz flicker, increased as task difficulty increased with number of regions; FPS players' frontal responses to the ignored 3 Hz flicker increased with number of regions. Thus the FPS players appear to employ an active suppression mechanism to deploy selective attention simultaneously to multiple interleaved regions, while RPG primarily use signal enhancement. These results suggest that fast-action gaming can affect neural strategies and the corresponding networks underlying attention, presumably by training mechanisms of distractor suppression.

The host proteins transportin SR2/TNPO3 and cyclophilin A exert opposing effects on HIV-1 uncoating.

Following entry of the HIV-1 core into target cells, productive infection depends on the proper disassembly of the viral capsid (uncoating). Although much is known regarding HIV-1 entry, the actions of host cell proteins that HIV-1 utilizes during early postentry steps are poorly understood. One such factor, transportin SR2 (TRN-SR2)/transportin 3 (TNPO3), promotes infection by HIV-1 and some other lentiviruses, and recent studies have genetically linked TNPO3 dependence of infection to the viral capsid protein (CA). Here we report that purified recombinant TNPO3 stimulates the uncoating of HIV-1 cores in vitro. The stimulatory effect was reduced by RanGTP, a known ligand for transportin family members. Depletion of TNPO3 in target cells rendered HIV-1 less susceptible to inhibition by PF74, a small-molecule HIV-1 inhibitor that induces premature uncoating. In contrast to the case for TNPO3, addition of the CA-binding host protein cyclophilin A (CypA) inhibited HIV-1 uncoating and reduced the stimulatory effect of TNPO3 on uncoating in vitro. In cells in which TNPO3 was depleted, HIV-1 infection was enhanced 4-fold by addition of cyclosporine, indicating that the requirement for TNPO3 in HIV-1 infection is modulated by CypA-CA interactions. Although TNPO3 was localized primarily to the cytoplasm, depletion of TNPO3 from target cells inhibited HIV-1 infection without reducing the accumulation of nuclear proviral DNA, suggesting that TNPO3 facilitates a stage of the virus life cycle subsequent to nuclear entry. Our results suggest that TNPO3 and cyclophilin A facilitate HIV-1 infection by coordinating proper uncoating of the core in target cells.

Retroviral integrase proteins and HIV-1 DNA integration.

Retroviral integrases catalyze two reactions, 3'-processing of viral DNA ends, followed by integration of the processed ends into chromosomal DNA. X-ray crystal structures of integrase-DNA complexes from prototype foamy virus, a member of the Spumavirus genus of Retroviridae, have revealed the structural basis of integration and how clinically relevant integrase strand transfer inhibitors work. Underscoring the translational potential of targeting virus-host interactions, small molecules that bind at the host factor lens epithelium-derived growth factor/p75-binding site on HIV-1 integrase promote dimerization and inhibit integrase-viral DNA assembly and catalysis. Here, we review recent advances in our knowledge of HIV-1 DNA integration, as well as future research directions.

Structural biology of retroviral DNA integration.

Three-dimensional macromolecular structures shed critical light on biological mechanism and facilitate development of small molecule inhibitors. Clinical success of raltegravir, a potent inhibitor of HIV-1 integrase, demonstrated the utility of this viral DNA recombinase as an antiviral target. A variety of partial integrase structures reported in the past 16 years have been instrumental and very informative to the field. Nonetheless, because integrase protein fragments are unable to functionally engage the viral DNA substrate critical for strand transfer inhibitor binding, the early structures did little to materially impact drug development efforts. However, recent results based on prototype foamy virus integrase have fully reversed this trend, as a number of X-ray crystal structures of active integrase-DNA complexes revealed key mechanistic details and moreover established the foundation of HIV-1 integrase strand transfer inhibitor action. In this review we discuss the landmarks in the progress of integrase structural biology during the past 17 years.

Structure-based modeling of the functional HIV-1 intasome and its inhibition.

The intasome is the basic recombination unit of retroviral integration, comprising the integrase protein and the ends of the viral DNA made by reverse transcription. Clinical inhibitors preferentially target the DNA-bound form of integrase as compared with the free protein, highlighting the critical requirement for detailed understanding of HIV-1 intasome structure and function. Although previous biochemical studies identified integrase residues that contact the DNA, structural details of protein-protein and protein-DNA interactions within the functional intasome were lacking. The recent crystal structure of the prototype foamy virus (PFV) integrase-viral DNA complex revealed numerous details of this related integration machine. Structures of drug-bound PFV intasomes moreover elucidated the mechanism of inhibitor action. Herein we present a model for the HIV-1 intasome assembled using the PFV structure as template. Our results pinpoint previously identified protein-DNA contacts within the quaternary structure and reveal hitherto unknown roles for Arg20 and Lys266 in DNA binding and integrase function. Models for clinical inhibitors bound at the HIV-1 integrase active site were also constructed and compared with previous studies. Our findings highlight the structural basis for HIV-1 integration and define the mechanism of its inhibition, which should help in formulating new drugs to inhibit viruses resistant to first-in-class compounds.

Cloning, expression and efficient refolding of carbohydrate-peptide mimicry recognizing single chain antibody 2D10.

Carbohydrate-peptide mimicry was found to be manifested through the cross-reactivity of an anti-mannopyranoside monoclonal antibody 2D10 (mAb-2D10) with YPY motif containing 12-mer peptide (DVFYPYPYASGS). Such multiple binding options for a monoclonal antibody could emanate from the possible flexibility of the antigen combining site. To address the molecular details of this phenomenon, single chain antibody (scFv) containing the antigen combining variable domain of mAb-2D10 was constructed. The present work describes the cloning, expression, purification and efficient refolding of scFv-2D10 and its His(6) tag fusion variants. The scFv expressed poorly in soluble/active form in the periplasmic compartment and concurrently exhibited higher tendency towards accumulation in inclusion bodies inside the Escherichia coli cytoplasm. The scFv was refolded from the inclusion bodies with approximately 68% yield using a previously described protocol which employed concomitant removal of the chaotropic and oxidizing reagents along with the additives. However, their differential removal, as described in the present report resulted in approximately 97% effective yield of the soluble scFv-2D10, an increase of 42%. The binding kinetics of the refolded scFv for both the mimicking ligands was examined using surface plasmon resonance experiments. The scFv-2D10 exhibited binding affinities similar to those reported for mAb-2D10 (IgG) showing that the modifications introduced in the refolding protocol have facilitated efficient preparation of active 2D10 scFv.

The requirement for cellular transportin 3 (TNPO3 or TRN-SR2) during infection maps to human immunodeficiency virus type 1 capsid and not integrase.

Recent genome-wide screens have highlighted an important role for transportin 3 in human immunodeficiency virus type 1 (HIV-1) infection and preintegration complex (PIC) nuclear import. Moreover, HIV-1 integrase interacted with recombinant transportin 3 protein under conditions whereby Moloney murine leukemia virus (MLV) integrase failed to do so, suggesting that integrase-transportin 3 interactions might underscore active retroviral PIC nuclear import. Here we correlate infectivity defects in transportin 3 knockdown cells with in vitro protein binding affinities for an expanded set of retroviruses that include simian immunodeficiency virus (SIV), bovine immunodeficiency virus (BIV), equine infectious anemia virus (EIAV), feline immunodeficiency virus (FIV), and Rous sarcoma virus (RSV) to critically address the role of integrase-transportin 3 interactions in viral infection. Lentiviruses, with the exception of FIV, display a requirement for transportin 3 in comparison to MLV and RSV, yielding an infection-based dependency ranking of SIV > HIV-1 > BIV and EIAV > MLV, RSV, and FIV. In vitro pulldown and surface plasmon resonance assays, in contrast, define a notably different integrase-transportin 3 binding hierarchy: FIV, HIV-1, and BIV > SIV and MLV > EIAV. Our results therefore fail to support a critical role for integrase binding in dictating transportin 3 dependency during retrovirus infection. In addition to integrase, capsid has been highlighted as a retroviral nuclear import determinant. Accordingly, MLV/HIV-1 chimera viruses pinpoint the genetic determinant of sensitization to transportin 3 knockdown to the HIV-1 capsid protein. We therefore conclude that capsid, not integrase, is the dominant viral factor that dictates transportin 3 dependency during HIV-1 infection.

Biochemical and virological analysis of the 18-residue C-terminal tail of HIV-1 integrase.

BACKGROUND: The 18 residue tail abutting the SH3 fold that comprises the heart of the C-terminal domain is the only part of HIV-1 integrase yet to be visualized by structural biology. To ascertain the role of the tail region in integrase function and HIV-1 replication, a set of deletion mutants that successively lacked three amino acids was constructed and analyzed in a variety of biochemical and virus infection assays. HIV-1/2 chimers, which harbored the analogous 23-mer HIV-2 tail in place of the HIV-1 sequence, were also studied. Because integrase mutations can affect steps in the replication cycle other than integration, defective mutant viruses were tested for integrase protein content and reverse transcription in addition to integration. The F185K core domain mutation, which increases integrase protein solubility, was furthermore analyzed in a subset of mutants. RESULTS: Purified proteins were assessed for in vitro levels of 3' processing and DNA strand transfer activities whereas HIV-1 infectivity was measured using luciferase reporter viruses. Deletions lacking up to 9 amino acids (1-285, 1-282, and 1-279) displayed near wild-type activities in vitro and during infection. Further deletion yielded two viruses, HIV-1(1-276) and HIV-1(1-273), that displayed approximately two and 5-fold infectivity defects, respectively, due to reduced integrase function. Deletion mutant HIV-1(1-270) and the HIV-1/2 chimera were non-infectious and displayed approximately 3 to 4-fold reverse transcription in addition to severe integration defects. Removal of four additional residues, which encompassed the C-terminal beta strand of the SH3 fold, further compromised integrase incorporation into virions and reverse transcription. CONCLUSION: HIV-1(1-270), HIV-1(1-266), and the HIV-1/2 chimera were typed as class II mutant viruses due to their pleiotropic replication defects. We speculate that residues 271-273 might play a role in mediating the known integrase-reverse transcriptase interaction, as their removal unveiled a reverse transcription defect. The F185K mutation reduced the in vitro activities of 1-279 and 1-276 integrases by about 25%. Mutant proteins 1-279/F185K and 1-276/F185K are therefore highlighted as potential structural biology candidates, whereas further deleted tail variants (1-273/F185K or 1-270/F185K) are less desirable due to marginal or undetectable levels of integrase function.

Role of antibody paratope conformational flexibility in the manifestation of molecular mimicry.

Molecular mimicry is a recurrent theme in host defense processes. The correlation of functional mimicry with the structural features of the antibody paratope has been investigated, addressing the consequences of mimicry in host immune mechanisms. Two anti-mannopyranoside antibodies, 1H7 and 2D10, representing the possible extremes of the recognition spectrum with regard to peptide-carbohydrate mimicry were examined. Crystallographic and molecular dynamics simulation analyses established correlation between the antibody flexibility and the manifestation of mimicry. It was evident that monoclonal antibody (mAb) 1H7, which has a narrow specificity in favor of the immunizing antigen, exhibited structural invariance. On the other hand, the antigen-combining site of 2D10, the mimicry-recognizing antibody, showed substantial divergence in the complementarity determining region loops. The docking of mannopyranoside within the antibody paratope revealed multiple modes of binding of the carbohydrate antigen in mAb 2D10 vis à vis single docking mode in mAb 1H7, which overlapped with the common monosaccharide binding site defined in anti-carbohydrate antibodies. The presence of additional antigen binding modes is perhaps reflective of the utilization of conformational flexibility in molecular mimicry. A relatively broader recognition repertoire--attributable to paratope flexibility--may facilitate the recognition of altered antigens of invading pathogens while the antibodies with narrow recognition specificity maintain the fidelity of the response.

Paratope plasticity in diverse modes facilitates molecular mimicry in antibody response.

The immune response against methyl-alpha-D-mannopyranoside mimicking 12-mer peptide (DVFYPYPYASGS) was analyzed at the molecular level towards understanding the equivalence of these otherwise disparate Ags. The Ab 7C4 recognized the immunizing peptide and its mimicking carbohydrate Ag with comparable affinities. Thermodynamic analyses of the binding interactions of both molecules suggested that the mAb 7C4 paratope lacks substantial conformational flexibility, an obvious possibility for facilitating binding to chemically dissimilar Ags. Favorable changes in entropy during binding indicated the importance of hydrophobic interactions in recognition of the mimicking carbohydrate Ag. Indeed, the topology of the Ag-combining site was dominated by a cluster of aromatic residues, contributed primarily by the specificity defining CDR H3. Epitope-mapping analysis demonstrated the critical role of three aromatic residues of the 12-mer in binding to the Ab. Our studies delineate a mechanism by which mimicry is manifested in the absence of either structural similarity of the epitopes or conformational flexibility in the paratope. An alternate mode of recognition of dissimilar yet mimicking Ags by the anti-peptide Ab involves plasticity associated with aromatic/hydrophobic and van der Waals interactions. Thus, antigenic mimicry may be a consequence of paratope-specific modulations rather than being dependent only on the properties of the epitope. Such modulations may have evolved toward minimizing the consequences of antigenic variation by invading pathogens.

A comparison of neural feature extraction methods for brain-machine interfaces.

Brain-machine interfaces (BMIs) have shown promise in augmenting people's control of their surroundings, especially for those suffering from paralysis due to neurological disorders. This paper describes an experiment using the rodent model to explore information available in neural signals recorded from chronically implanted intracortical microelectrode arrays. In offline experiments, a number of neural feature extraction methods were utilized to obtain neural activity vectors (NAVs) describing the activity of the underlying neural population while rats performed a discrimination task. The methods evaluated included standard techniques such as binned spike rates and local field potential spectra as well as more novel approaches including matched-filter energy, raw signal spectra, and an autocorrelation energy measure (AEM) approach. Support vector machines (SVMs) were trained offline to classify left from right going movements by utilizing features contained in the NAVs obtained by the different methods. Each method was evaluated for accuracy and robustness. Results show that most algorithms worked well for decoding neural signals both during and prior to movement, with spectral methods providing the best stability.

Plasticity within the antigen-combining site may manifest as molecular mimicry in the humoral immune response.

Structural and physiological facets of carbohydrate-peptide mimicry were addressed by analyzing the Ab response to alpha-d-mannopyranoside. mAbs against alpha-d-mannopyranoside were generated and screened with the carbohydrate-mimicking 12 mer (DVFYPYPYASGS) peptide. Three mAbs, 2D10, 1H11, and 1H7, which were subjected to detailed analysis, exhibit diverse V gene usage, indicating their independent germline origins. Although the mAb 1H7 was specific in binding only to the immunizing Ag, the Abs 2D10 and 1H11 recognize the 12 mer peptide as well as the immunogen, alpha-d-mannopyranoside. The Abs that recognize mimicry appear to bind to a common epitope on the peptide and do not share the mode of peptide binding with Con A. Binding kinetics and thermodynamics of Ag recognition suggest that the Ab that does not recognize peptide-carbohydrate mimicry probably has a predesigned mannopyranoside-complementing site. In contrast, the mimicry-recognizing Abs adopt the Ag-combining site only on exposure to the sugar, exploiting the conformational flexibility in the CDRs. Although the mAb 1H7 showed unique specificity toward mannopyranoside, the mimicry-recognizing Abs 2D10 and 1H11 exhibited degenerate specificities with regard to other sugar moieties. It is proposed that the degeneracy of specificity arising from the plasticity at the Ag-combining site in a subset of the Ab clones may be responsible for exhibiting molecular mimicry in the context of Ab response.