Rachita Dash

Talanta. 2024 Jun 22;278:126458. doi: 10.1016/j.talanta.2024.126458. Online ahead of print.

ABSTRACT

A modified development protocol and concomitant characterisation of a first generation biosensor for the detection of brain extracellular d-serine is reported. Functional parameters important for neurochemical monitoring, including sensor sensitivity, O₂ interference, selectivity, shelf-life and biocompatibility were examined. Construction and development involved the enzyme d-amino acid oxidase (DAAO), utilising a dip-coating immobilisation method employing a new extended drying approach. The resultant Pt-based polymer enzyme composite sensor achieved high sensitivity to d-serine (0.76 ± 0.04 nA mm^-2. μM^-1) and a low μM limit of detection (0.33 ± 0.02 μM). The in-vitro response time was within the solution stirring time, suggesting potential sub-second in-vivo response characteristics. Oxygen interference studies demonstrated a 1 % reduction in current at 50 μM O₂ when compared to atmospheric O₂ levels (200 μM), indicating that the sensor can be used for reliable neurochemical monitoring of d-serine, free from changes in current associated with physiological O₂ fluctuations. Potential interference signals generated by the principal electroactive analytes present in the brain were minimised by using a permselective layer of poly(o-phenylenediamine), and although several d-amino acids are possible substrates for DAAO, their physiologically relevant signals were small relative to that for d-serine. Additionally, changing both temperature and pH over possible in vivo ranges (34-40 °C and 7.2-7.6 respectively) resulted in no significant effect on performance. Finally, the biosensor was implanted in the striatum of freely moving rats and used to monitor physiological changes in d-serine over a two-week period.

PMID:38955102 | DOI:10.1016/j.talanta.2024.126458

Chemistry. 2024 Jul 2:e202401654. doi: 10.1002/chem.202401654. Online ahead of print.

ABSTRACT

Cyclisation of peptides by forming thioether (lanthionine), disulfide (cystine) or methylene thioacetal bridges between side chains is established as an important tool to stabilise a given structure, enhance metabolic stability and optimise both potency and selectivity. However, a systematic comparative study of the effects of differing bridging modalities on peptide conformation has not previously been carried out. In this paper, we have used the NMR deconvolution algorithm, NAMFIS, to determine the conformational ensembles, in aqueous solution, of three cyclic analogues of angiotensin(1-7), incorporating either disulfide, or non-reduceable thioether or methylene thioacetal bridges. We demonstrate that the major solution conformations are conserved between the different bridged peptides, but the distribution of conformations differs appreciably. This suggests that subtle differences in ring size and bridging structure can be exploited to fine-tune the conformational properties of cyclic peptides, which may modulate their bioactivities.

PMID:38953277 | DOI:10.1002/chem.202401654

RSC Med Chem. 2024 Apr 19;15(6):2018-2029. doi: 10.1039/d4md00073k. eCollection 2024 Jun 19.

ABSTRACT

In a recent paper in this journal (RSC Med. Chem., 2023, 14, 2429), we described an unusually strong impact of regiospecific exchange of phenylalanines by tyrosines in 10 gallium-68-labeled trimers of certain cyclic RGD peptides, c[XRGDLAXp(NMe)K] (X = F or Y), on non-specific organ uptakes. We found that there was, in part, no correlation of liver uptake with established polarity proxies, such as the octanol-water distribution coefficient (log D). Since this observation could not be explained straightforwardly, we suggested that the symmetry of the compounds had resulted in a synergistic interaction of certain components of the macromolecules. In the present work, we investigated whether a comparable effect also occurred for a series of 5 tetramers labeled with lutetium-177. We found that in contrast to the trimers, liver uptake of the tetramers was well correlated to their polarity, indicating that the unusual observations along the trimer series indeed was a unique feature, probably related to their particular symmetry. Since the Lu-177 labeled tetramers are also potential agents for treatment of a variety of αvβ6-integrin expressing cancers, these were evaluated in mice bearing human lung adenocarcinoma xenografts. Due to their tumor-specific uptake and retention in biodistribution and SPECT imaging experiments, these compounds are considered a step forward on the way to αvβ6-integrin-targeted anticancer agents. Furthermore, we noticed that the presence of tyrosines in general had a positive impact on the in vivo performance of our peptide multimers. In view of the fact that a corresponding rule was already proposed in the context of protein engineering, we argue in favor of considering peptide multimers as a special class of small or medium-sized proteins. In summary, we contend that the performance of peptide multimers is less determined by the in vitro characteristics (particularly, affinity and selectivity) of monomers, but rather by the peptides' suitability for the overall macromolecular design concept, and peptides containing tyrosines are preferred.

PMID:38911160 | PMC:PMC11187563 | DOI:10.1039/d4md00073k

bioRxiv [Preprint]. 2024 Jun 16:2024.06.15.590358. doi: 10.1101/2024.06.15.590358.

ABSTRACT

Proteins composed of a single structural unit tandemly repeated multiple times carry out a wide range of functions in biology. There has hence been considerable interest in designing such repeat proteins; previous approaches have employed strict constraints on secondary structure types and relative geometries, and most characterized designs either mimic a known natural topology, adhere closely to a parametric helical bundle architecture, or exploit very short repetitive sequences. Here, we describe Rosetta-based and deep learning hallucination methods for generating novel repeat protein architectures featuring mixed alpha-helix and beta-strand topologies, and 25 new highly stable alpha-beta proteins designed using these methods. We find that incorporation of terminal caps which prevent beta strand mediated intermolecular interactions increases the solubility and monomericity of individual designs as well as overall design success rate.

PMID:38915539 | PMC:PMC11195203 | DOI:10.1101/2024.06.15.590358

J Am Chem Soc. 2024 Jul 3;146(26):17691-17699. doi: 10.1021/jacs.4c01979. Epub 2024 Jun 18.

ABSTRACT

Nonproteinogenic amino acids, including d-α-, β-, and γ-amino acids, present in bioactive peptides play pivotal roles in their biochemical activities and proteolytic stabilities. d-α-Amino acids (dαAA) are widely used building blocks that can enhance the proteolytic stability. Cyclic β^2,3-amino acids (cβAA), for instance, can fold peptides into rigid secondary structures, improving the binding affinity and proteolytic stability. Cyclic γ^2,4-amino acids (cγAA) are recently highlighted as rigid residues capable of preventing the proteolysis of flanking residues. Simultaneous incorporation of all dαAA, cβAA, and cγAA into a peptide is expected to yield l-α/d-α/β/γ-hybrid peptides with improved stability and potency. Despite challenges in the ribosomal incorporation of multiple nonproteinogenic amino acids, our engineered tRNA^Pro1E2 successfully reaches such a difficulty. Here, we report the ribosomal synthesis of macrocyclic l-α/d-α/β/γ-hybrid peptide libraries and their application to in vitro selection against interferon gamma receptor 1 (IFNGR1). One of the resulting l-α/d-α/β/γ-hybrid peptides, IB1, exhibited remarkable inhibitory activity against the IFN-γ/IFNGR1 protein-protein interaction (PPI) (IC₅₀ = 12 nM), primarily attributed to the presence of a cβAA in the sequence. Additionally, cγAAs and dαAAs in the resulting peptides contributed to their serum stability. Furthermore, our peptides effectively inhibit IFN-γ/IFNGR1 PPI at the cellular level (best IC₅₀ = 0.75 μM). Altogether, our platform expands the chemical space available for exploring peptides with high activity and stability, thereby enhancing their potential for drug discovery.

PMID:38888290 | PMC:PMC11229689 | DOI:10.1021/jacs.4c01979