Dissecting dual specificity: Identifying key residues in L-asparaginase for enhanced acute lymphoid leukemia therapy and reduced adverse effects.

L-asparaginase from Escherichia coli (EcA) has been used for the treatment of acute lymphoid leukemia (ALL) since the 1970s. Nevertheless, the enzyme has a second specificity that results in glutaminase breakdown, resulting in depletion from the patient's body, causing severe adverse effects. Despite the huge interest in the use of this enzyme, the exact process of glutamine depletion is still unknown and there is no consensus regarding L-asparagine hydrolysis. Here, we investigate the role of T12, Y25, and T89 in asparaginase and glutaminase activities. We obtained individual clones containing mutations in the T12, Y25 or T89 residues. After the recombinant production of wild-type and mutated EcA, The purified samples were subjected to structural analysis using Nano Differential Scanning Fluorimetry, which revealed that all samples contained thermostable molecules in their active structural conformation, the homotetramer conformation. The quaternary conformation was confirmed by DLS and SEC. The activity enzymatic assay combined with molecular dynamics simulation identified the contribution of T12, Y25, and T89 residues in EcA glutaminase and asparaginase activities. Our results mapped the enzymatic behavior paving the way for the designing of improved EcA enzymes, which is important in the treatment of ALL.

Di-n-butyl phthalate negatively affects humic acid conversion and microbial enzymatic dynamics during composting.

To understand the effects of phthalic acid esters (PAEs) on humic acid (HA) conversion, enzymatic and specific metabolic dynamics during composting under di-n-butyl phthalate (DBP) stress were evaluated for the first time. The results indicated that HA conversion was mainly related to bacteria rather than fungi, with positive associations with Actinobacteria, Chloroflexi, and Gemmatimonadota (all P < 0.05), and negative associations with Proteobacteria and Bacteroidota (all P < 0.05), while DBP stress retarded HA formation by altering the core microbes related to HA formation and their metabolic functions. Moreover, typical hydrolase and oxidoreductase activities were altered under DBP stress, proteases and cellulases were hindered, and peroxidases as well as polyphenol oxidases were promoted during composting. Overall, our data shows that DBP stress can retard HA formation and compost maturation by interfering with microbial activity. This study provides potentially useful information for the degradation and reuse of PAE-contaminated waste.

Study on Inhibitory Effects of Ethanol Extract of Different Medicinal Parts from Syzygium jambos on the Activities of α- Glycosidase and α-Amylase / 中国药房

OBJECTIVE： To compare inhibitory effects of ethanol extract of different medicinal parts （root， stem， leaf， seed， flower and flesh） from Syzygium jambos on the activities of α-glycosidase and α-amylase. METHODS： Using half-inhibitory concentration value （IC50） as evaluation index， acarbose as positive control， inhibitory effects of ethanol extract of different medicinal parts from S. jambos on the activities of α-glycosidase （from yeast and small instestine in mice） and α-amylase were evaluated with in vitro inhibition model. The enzymatic dynamics and Lineweaver-Burk methods were used to analyze the inhibitory type of the best medicinal part on the activities of α-glycosidase and α-amylase. RESULTS： In the yeast α-glucosidase inhibitory activity test， the order of inhibitory activity was S. jambos seed＞S. jambos stem＞S. jambos leaf＞S. jambos root＞S. jambos flower＞S. jambos flesh＞acarbose. In the mice intestine α-glucosidase inhibitory activity test， the order of inhibitory activity was S. jambos seed＞S. jambos stem＞S. jambos root＞S. jambos leaf＞S. jambos flower＞S. jambos flesh＞acarbose. In the α-amylase inhibitory activity test， the order of inhibitory activity was acarbose＞S. jambos seed＞S. jambos stem＞S. jambos root＞S. jambos leaf＞S. jambos flesh＞S. jambos flower. Ethanol extract of S. jambos seed had the stronger inhibition activity against α-glucosidase from yeast，α-glucosidase from small intestine in mice and α-amylase than other medicinal parts [IC50 were（6.64±0.24）， （32.77±2.46） and （41.18±1.63） μg/mL]. Ethanol extract of S. jambos seed had the stronger inhibition activity against α-glucosidase than acarbose [IC50 to α-glucosidase from yeast and α-glucosidase from small intestine in mice were （2 833.33±5.48）， （1 304.21±6.45） μg/mL] （P＜0.05）. The inhibitory effect of ethanol extract from S. jambos on the activity of α-amylase was less than that of acarbose [IC50 was （27.27±1.24） μg/mL] （P＜0.05）. Enzymatic dynamics showed that the inhibitory type of ethanol extract from S. jambos seed on α-glucosidase and α-amylase were both reversible competitive inhibition. CONCLUSIONS： Among different parts of S. jambos such as root， stem， leaf， seed， flower and flesh， S. jambos seed shows the strongest inhibitory effects on the activities of α-glucosidase and α-amylase， which has the value of being developed for the treatment of diabetes or health food.