INTRODUCTION: Imprinting broadly neutralizing antibody (bNAb) paratopes by shape complementary protein mimotopes represents a potential alternative for developing vaccine immunogens. This approach, designated as a Non-Cognate Ligand Strategy (NCLS), has recently been used for the identification of protein variants mimicking CD4 binding region epitope or membrane proximal external region (MPER) epitope of HIV-1 envelope (Env) glycoprotein. However, the potential of small binding proteins to mimic viral glycan-containing epitopes has not yet been verified. METHODS: In this work, we employed a highly complex combinatorial Myomedin scaffold library to identify variants recognizing paratopes of super candidate bNAbs, PGT121 and PGT126, specific for HIV-1 V3 loop epitopes. RESULTS: In the collection of Myomedins called MLD variants targeted to PGT121, three candidates competed with gp120 for binding to this bNAb in ELISA, thus suggesting an overlapping binding site and epitope-mimicking potential. Myomedins targeted to PGT126 designated MLB also provided variants that competed with gp120. Immunization of mice with MLB or MLD binders resulted in the production of anti-gp120 and -Env serum antibodies. Mouse hyper-immune sera elicited with MLB036, MLB041, MLB049, and MLD108 moderately neutralized 8-to-10 of 22 tested HIV-1-pseudotyped viruses of A, B, and C clades in vitro. DISCUSSION: Our data demonstrate that Myomedin-derived variants can mimic particular V3 glycan epitopes of prominent anti-HIV-1 bNAbs, ascertain the potential of particular glycans controlling neutralizing sensitivity of individual HIV-1 pseudoviruses, and represent promising prophylactic candidates for HIV-1 vaccine development.
Proteins are naturally formed by domains edging their functional and structural properties. A domain out of the context of an entire protein can retain its structure and to some extent also function on its own. These properties rationalize construction of artificial fusion multidomain proteins with unique combination of various functions. Information on the specific functional and structural characteristics of individual domains in the context of new artificial fusion proteins is inevitably encoded in sequential order of composing domains defining their mutual spatial positions. So the challenges in designing new proteins with new domain combinations lie dominantly in structure/function prediction and its context dependency. Despite the enormous body of publications on artificial fusion proteins, the task of their structure/function prediction is complex and nontrivial. The degree of spatial freedom facilitated by a linker between domains and their mutual orientation driven by noncovalent interactions is beyond a simple and straightforward methodology to predict their structure with reasonable accuracy. In the presented manuscript, we tested methodology using available modeling tools and computational methods. We show that the process and methodology of such prediction are not straightforward and must be done with care even when recently introduced AlphaFold II is used. We also addressed a question of benchmarking standards for prediction of multidomain protein structures-x-ray or Nuclear Magnetic Resonance experiments. On the study of six two-domain protein chimeras as well as their composing domains and their x-ray structures selected from PDB, we conclude that the major obstacle for justified prediction is inappropriate sampling of the conformational space by the explored methods. On the other hands, we can still address particular steps of the methodology and improve the process of chimera proteins prediction.
Úvod: Moderní diagnostika urolitiázy v dnešní době využívá především spirálního CT vyšetření. Při vyšetření je možné kromě běžných morfologických dat zjistit také denzitu konkrementu. Práce se zabývá možnou korelací mezi denzitou konkrementu a jeho chemickým složením. Materiál a metoda: Autoři prezentují retrospektivní zhodnocení výsledků chemického složení konkrementů u 154 pacientů ze dvou pracovišť, kteří podstoupili operační léčbu urolitiázy nebo konkrementy spontánně vymočili. U všech pacientů jsme získali data denzity konkrementů z CT vyšetření provedených v době diagnostiky urolitiázy nebo v průběhu kontrolních vyšetření. Výsledky: V uvedeném souboru se nám podařilo prokázat statistický významný rozdíl mezi denzitou urátových a kalciových konkrementů (p < 0,05). Rozdíl denzit mezi ostatními skupinami konkrementů nebyl statisticky významný. Korelaci mezi denzitou konkrementu a jeho chemickým složením jsme neprokázali. Závěr: Použití nativního spirálního CT vyšetření v diagnostice urolitiázy umožňuje odlišit urátové konkrementy od kalciových. Predikce chemického složení konkrementu je důležitým faktorem při volbě optimální léčebné strategie.
Introduction: Today's modern methods of diagnosing urolithiasis mainly utilize spiral CT examination. In addition to common morphological data, it is possible to determine stone density by means of the examination. The paper deals with a possible correlation between stone density and its chemical composition. Material and method: The authors present a retrospective review of outcomes of the chemical composition of stones in 154 patients from two centres who underwent surgical treatment for urolithiasis or spontaneously passed stones in their urine. In all patients, data on stone density were obtained from CT examinations performed at the time of diagnosis of urolithiasis or during follow-up evaluations. Results: In the reported group, a statistically significant difference was shown between the density of urate and calcium stones (p < 0.05). The differences in densities between the other groups of stones were not statistically significant. A correlation between stone density and its chemical composition failed to be confirmed. Conclusion: The use of non-contrast spiral CT in diagnosing urolithiasis allows to distinguish between urate and calcium stones. Predicting the chemical composition of a stone is an important factor in selecting an optimal therapeutic strategy.
