Modifications to functional and biological properties of proteins of cowpea pulse crop by ultrasound-assisted extraction

Natural resource depletion, negative environmental effects and the challenge to secure global food security led to the establishment of the Sustainable Development Goals (SDGs). In need to explore underutilized sustainable protein sources, this study aims at isolating protein from cowpea by ultrasound-assisted extraction (UAE), where the techno-functional characteristics of the protein isolates were studied at different sonication conditions i.e., 100 W and 200 W at processing times ranging from 5 to 20 min. The US at 200 W-10 min produced the optimal results for all properties. In this process combination, there was an increase in protein yield, solubility, water-holding capacity, foaming capacity and stability, emulsion activity and stability, zeta-potential, and in-vitro protein digestibility from 31.78% to 58.96%, 57.26% to 68.85%, 3.06 g/g to 3.68 g/g 70.64% to 83.74%, 30.76% to 60.01%, 47.48% to 64.26%, 56.59% to 87.71%, –32.9 mV to −44.2 mV and 88.27% to 89.99%, respectively and particle size dropped from 763 nm to 559 nm in comparison to control. The microstructure and secondary-structure alterations of proteins caused by sonication were validated by SEM images, SDS-PAGE, and FTIR analyses. Sonication leads to acoustic cavitation and penetrate the cell walls, improving extraction from the solid to liquid phase. After sonication, the hydrophobic protein groups were exposed and proteins were partially denatured which increased its functionality. The findings demonstrated that UAE of cowpea protein improved yield, modify characteristics to fit the needs of the food industry, and contribute to achieving SDGs 2, 3, 7, 12, and 13.     

Viscosity of concentrated protein solutions

The viscosity of solutions of four proteins (Bovine Serum Albumin, Ovalbumin, _s-1 Casein, Lysozyme), brought to the random coil conformation, has been measured over a large concentration range extending into the entanglement region. A master curve is obtained in the dilute and semi-dilute regions with the reduced variables and of Simha and Utracki.By using Graessley's expression for the polymer coil expansion at a given concentration in the semi-dilute region (c ^* c c ^**), a simple equation is established giving the relative viscosity _r as a function of concentrationc: forc ^* c c ^**, ln _r = 2a[]c ^*(c/c ^*)^1/2a – (2a - 1)[]c ^*; wherec ^* is the incipient overlap concentration, [] the intrinsic viscosity, anda the Mark-Houwink exponent for the polymer-solvent considered.This equation fits well the experimental results. The adjustment yields for the parametera values which are comprised between 0.6 and 0.7, as expected, for Bovine Serum Albumin and Ovalbumin, but very close to 0.5 for _s-1 Casein and Lysozyme. This can be explained by the fact that the molecular weights of the two latter proteins are lower than, or very close, the critical molecular weight; the critical molecular weight is estimated to be about 20000.     

Electronic Olfactory Sensor Based on A. mellifera Odorant‐Binding Protein 14 on a Reduced Graphene Oxide Field‐Effect Transistor

An olfactory biosensor based on a reduced graphene oxide (rGO) field‐effect transistor (FET), functionalized by the odorant‐binding protein 14 (OBP14) from the honey bee (Apis mellifera) has been designed for the in situ and real‐time monitoring of a broad spectrum of odorants in aqueous solutions known to be attractants for bees. The electrical measurements of the binding of all tested odorants are shown to follow the Langmuir model for ligand–receptor interactions. The results demonstrate that OBP14 is able to bind odorants even after immobilization on rGO and can discriminate between ligands binding within a range of dissociation constants from K_d=4 μM to K_d=3.3 mM . The strongest ligands, such as homovanillic acid, eugenol, and methyl vanillate all contain a hydroxy group which is apparently important for the strong interaction with the protein.     

Structure and Mechanism of an Aspartimide‐Dependent Peptide Ligase in Human Legumain

Peptide ligases expand the repertoire of genetically encoded protein architectures by synthesizing new peptide bonds, energetically driven by ATP or NTPs. Here, we report the discovery of a genuine ligase activity in human legumain (AEP) which has important roles in immunity and tumor progression that were believed to be due to its established cysteine protease activity. Defying dogma, the ligase reaction is independent of the catalytic cysteine but exploits an endogenous energy reservoir that results from the conversion of a conserved aspartate to a metastable aspartimide. Legumain’s dual protease–ligase activities are pH‐ and thus localization controlled, dominating at acidic and neutral pH, respectively. Their relevance includes reversible on–off switching of cystatin inhibitors and enzyme (in)activation, and may affect the generation of three‐dimensional MHC epitopes. The aspartate–aspartimide (succinimide) pair represents a new paradigm of coupling endergonic reactions in ATP‐scarce environments.     

Engineering Rieske Non‐Heme Iron Oxygenases for the Asymmetric Dihydroxylation of Alkenes

The asymmetric dihydroxylation of olefins is of special interest due to the facile transformation of the chiral diol products into valuable derivatives. Rieske non‐heme iron oxygenases (ROs) represent promising biocatalysts for this reaction as they can be engineered to efficiently catalyze the selective mono‐ and dihydroxylation of various olefins. The introduction of a single point mutation improved selectivities (≥95 %) and conversions (>99 %) towards selected alkenes. By modifying the size of one active site amino acid side chain, we were able to modulate the regio‐ and stereoselectivity of these enzymes. For distinct substrates, mutants displayed altered regioselectivities or even favored opposite enantiomers compared to the wild‐type ROs, offering a sustainable approach for the oxyfunctionalization of a wide variety of structurally different olefins.     

Oriented Protein Nanoarrays on Block Copolymer Template

Here, a simple yet robust method is developed to fabricate oriented protein nanoarrays by employing a block copolymer (BCP) template, which presents nano‐scaled spot areas at high‐density arrays. Unlike the conventional BCP nanolithography, the BCP platform described here resists nonspecific protein adsorption and prevents the denaturation of immobilized proteins in aqueous solution. The orderly arranged array areas are functionalized by linking chemistry which allows for the precise control of protein orientation. This approach allows us to generate potentially oriented protein nanoarrays at high‐density array spots, which is useful for miniaturized nanoarrays within high‐throughput proteomic applications.

     

Selenium and Selenocysteine in Protein Chemistry

Selenocysteine, the selenium‐containing analogue of cysteine, is the twenty‐first proteinogenic amino acid. Since its discovery almost fifty years ago, it has been exploited in unnatural systems even more often than in natural systems. Selenocysteine chemistry has attracted the attention of many chemists in the field of chemical biology owing to its high reactivity and resulting potential for various applications such as chemical modification, chemical protein (semi)synthesis, and protein folding, to name a few. In this Minireview, we will focus on the chemistry of selenium and selenocysteine and their utility in protein chemistry.     

Unambiguous Determination of Protein Arginine Ionization States in Solution by NMR Spectroscopy

Because arginine residues in proteins are expected to be in their protonated form almost without exception, reports demonstrating that a protein arginine residue is charge‐neutral are rare and potentially controversial. Herein, we present a ¹³C‐detected NMR experiment for probing individual arginine residues in proteins notwithstanding the presence of chemical and conformational exchange effects. In the experiment, the ¹⁵N^η and ¹⁵N^ϵ chemical shifts of an arginine head group are correlated with that of the directly attached ¹³C^ζ. In the resulting spectrum, the number of protons in the arginine head group can be obtained directly from the ¹⁵N–¹H scalar coupling splitting pattern. We applied this method to unambiguously determine the ionization state of the R52 side chain in the photoactive yellow protein from Halorhodospira halophila . Although only three H^η atoms were previously identified by neutron crystallography, we show that R52 is predominantly protonated in solution.