Thermophilic PHP Protein Tyrosine Phosphatases (Cap8C and Wzb) from Mesophilic Bacteria.

Protein tyrosine phosphatases (PTPs) of the polymerase and histidinol phosphatase (PHP) superfamily with characteristic phosphatase activity dependent on divalent metal ions are found in many Gram-positive bacteria. Although members of this family are co-purified with metal ions, they still require the exogenous supply of metal ions for full activation. However, the specific roles these metal ions play during catalysis are yet to be well understood. Here, we report the metal ion requirement for phosphatase activities of S. aureus Cap8C and L. rhamnosus Wzb. AlphaFold-predicted structures of the two PTPs suggest that they are members of the PHP family. Like other PHP phosphatases, the two enzymes have a catalytic preference for Mn2+, Co2+ and Ni2+ ions. Cap8C and Wzb show an unusual thermophilic property with optimum activities over 75 °C. Consistent with this model, the activity-temperature profiles of the two enzymes are dependent on the divalent metal ion activating the enzyme.

Role of the C-Terminal ß Sandwich of Thermoanaerobacter tengcongensis Thermophilic Esterase in Hydrolysis of Long-Chain Acyl Substrates.

To search for a novel thermostable esterase for optimized industrial applications, esterase from a thermophilic eubacterium species, Thermoanaerobacter tengcongensis MB4, was purified and characterized in this work. Sequence analysis of T. tengcongensis esterase with other homologous esterases of the same family revealed an apparent tail at the C-terminal that is not conserved across the esterase family. Hence, it was hypothesized that the tail is unlikely to have an essential structural or catalytic role. However, there is no documented report of any role for this tail region. We probed the role of the C-terminal domain on the catalytic activity and substrate preference of T. tengcongensis esterase EstA3 with a view to see how it could be engineered for enhanced properties. To achieve this, we cloned, expressed, and purified the wild-type and the truncated versions of the enzyme. In addition, a naturally occurring member of the family (from Brevibacillus brevis) that lacks the C-terminal tail was also made. In vitro characterization of the purified enzymes showed that the C-terminal domain contributes significantly to the catalytic activity and distinct substrate preference of T. tengcongensis esterase EstA3. All three recombinant enzymes showed the highest preference for paranitrophenyl butyrate (pNPC4), which suggests they are true esterases, not lipases. Kinetic data revealed that truncation had a slight effect on the substrate-binding affinity. Thus, the drop in preference towards long-chain substrates might not be a result of substrate binding affinity alone. The findings from this work could form the basis for future protein engineering allowing the modification of esterase catalytic properties through domain swapping or by attaching a modular protein domain.

A review of some common African spices with antihypertensive potential.

Hypertension is the most common non-communicable disease, with about 1.28 billion hypertensive people worldwide. It is more prevalent in men than women and more common in the elderly. Hereditary, age, obesity, lifestyle, diet, alcohol, and chronic metabolic diseases are the major risk factors of hypertension. Treating hypertension is a complex process as there are several mechanisms responsible for its pathogenesis; hence, a combination of several drugs is used for managing hypertension. Drugs used in managing hypertension are expensive and often come with associated side effects; thus, there is need for alternative means of managing this life-threatening disease. These drugs do not achieve the recommended blood pressure target in most people; more so majority of people with hypertension do not follow the treatment regimen religiously. Some Africans have been reported to become normotensive as a result of dietary consumption of spices. Several spices have been used over the years in Africa to manage hypertension. The aim of this review is to evaluate the ethnomedicinal use, bioactive phytochemical composition, bioactive compounds present, and pharmacological applications of spices commonly used in Africa for managing hypertension. Most of the plants used contained polyphenols, flavonoids, tannins, anthraquinone, flavonoids, cardiac glycosides, and saponins. Dietary supplementation of Xylopia aethiopica and other spices in diet have been proven to significantly reduced plasma angiotensin-I-converting enzyme (ACE) than simvastatin (the reference drug). Toxicological, histological, and hematological evaluation revealed that acute and chronic consumption of most of these spices are safe. Studies have also revealed that some of the spices can be used as alternative therapy alongside usual antihypertensive medications. PRACTICAL IMPLICATION: The prevalent rate of hypertension is on the increase in both the developed and developing countries. People often skip medication due to their busy schedule and anti-hypertensive potential side effects; however, this is not the case with food/spices as most people consumed them daily. Deliberate, right combinations and consistent incorporation of spices with proven anti-hypertensive potential into our diet may be of great benefit in normalizing blood pressure and mitigate other complications on the heart and vital organs.

Kinetics of angiotensin -1 converting enzyme inhibition and antioxidative properties of Azadirachta indica seed protein hydrolysates.

Neem (Azadirachta indica) seed protein hydrolysates were investigated for in vitro antioxidant and angiotensin 1-converting enzyme (ACE)-inhibitory activities. Neem seed proteins were hydrolysed using pepsin, trypsin and Alcalase. The degree of pepsin hydrolysis of neem seed protein was significantly higher (p < 0.05) than those of trypsin and Alcalase hydrolysis. Proteolytic hydrolysis of the isolate resulted in hydrolysates with improved Arg/Lys ratio, with pepsin hydrolysates still being able to maintain an acceptable level of essential amino acids comparable to that of the isolate. At 2.5 mg/mL, pepsin neem seed protein hydrolysate (NSPH) demonstrated the strongest antioxidant activity with 67.15 % and 50.07 % DPPH- and superoxide anion radical-scavenging activities, respectively, while trypsin NSPH had the highest ferric-reducing power. Using N-[3-(2-furyl)acryloyl]-L-phenylalanyl-glycyl-glycine (FAPGG) as substrate, NSPHs strongly inhibited ACE (69.20-80.39 %) in a concentration-dependent manner. Pepsin NSPH had higher ACE-inhibitory activity than trypsin and Alcalase NSPHs. Kinetic studies showed the mechanism of ACE inhibition to be mixed-type with Ki values of 0.62, 0.84, 1.5 for pepsin, trypsin and alcalase NSPH, respectively. These results suggest that NSPH can be used as a potential nutraceutical with antioxidant capacity and inhibitory activity against ACE.

Distinct metal ion requirements for the phosphomonoesterase and phosphodiesterase activities of calf intestinal alkaline phosphatase.

The roles of Mg(2+) and Zn(2+) ions in promoting phosphoryl transfer catalysed by alkaline phosphatase are yet to be fully characterised. We investigated the divalent metal ion requirements for the monoesterase and diesterase activities of calf intestinal alkaline phosphatase. The synergistic effect of Mg(2+) and Zn(2+) in promoting the hydrolysis of para-nitrophenyl phosphate (monoesterase reaction) by alkaline phosphatase is not observed in the hydrolysis of the diesterase substrate, bis-para-nitrophenyl phosphate. Indeed, the diesterase reaction is inhibited by concentrations of Mg(2+) that were optimal for the monoesterase reaction. This study reveals that the substrate specificities of alkaline phosphatases and related bimetalloenzymes are subject to regulation by changes in the nature and availability of cofactors, and the different cofactor requirements of the monoesterase and diesterase reactions of mammalian alkaline phosphatases could have significance for the biological functions of the enzymes.