ABSTRACT
Organosulfur compounds are the basis for the unique aroma of Lentinula edodes, and cysteine sulfoxide lyase (C-S lyase) is the key enzyme in this trait. The enzyme from Alliium sativum has been crystallized and well-characterized; however, there have been no reports of the characterization of fungi C-S lyase at the molecular level. We identified a L. edodes C-S lyase (Lecsl), cloned a gene of Csl encoded Lecsl and then combined modeling, simulations, and experiments to understand the molecular basis of the function of Lecsl. Our analysis revealed Lecsl to be a novel cysteine desulfurase and not a type of cysteine sulfoxide lyase. The pyridoxal-5-phosphate (PLP) molecule bonded tightly to Lecsl to form a Lecsl-PLP complex. Moreover, the Lecsl had one active center that served to bind two kinds of substrates, S-methyl-L-cysteine sulfoxide and L-cysteine, and had both cysteine sulfoxide lyase and cysteine desulfurase activity. We found that the amino acid residue Asn393 was essential for the catalytic activity of Lecsl and that the gene Csl encoded a novel cysteine desulfurase to influence organosulfur compounds in L. edodes. Our results provide a new insight into understanding the formation of the unique aroma of L. edodes.
Subject(s)
Carbon-Sulfur Lyases/metabolism , Shiitake Mushrooms/metabolism , Sulfur Compounds/metabolism , Amino Acid Sequence , Lyases/metabolism , Molecular Sequence Data , Pyridoxal Phosphate/metabolism , Substrate SpecificityABSTRACT
OBJECTIVE: This study aimed to evaluate the diagnostic value of a 24-hour esophageal combined multichannel intraluminal impedance pH (24â h our MII-pH) monitoring in children with gastroesophageal reflux (GER) disease and recurrent pneumonia. METHODS: A total of 17 cases with a suspected diagnosis of GER disease children with unexplained recurrent pneumonia underwent a 24-hour MII-pH monitoring to analysis of the nature and characteristics of reflux. RESULTS: In the 17 cases of GER disease with recurrent pneumonia, 11 (65%) were confirmed positively by a 24-hours of MII-pH monitoring. A total of 853 reflux cycle were detected, of which 65.3% were acid refluxes predominantly occurring within 2 hours after meal, and 71.6% were mixed refluxes. Refluxes occurred most frequently in the supine position, followed by the lateral position and the prone position. Distal reflux was the predominant form of reflux. According to the Biox-Ochoa classification, 73% of cases (8/11) were severe GER. CONCLUSIONS: MII-pH monitoring may effectively characterize refluxate properties and thereby increase the detection rate of GER disease.
Subject(s)
Gastroesophageal Reflux/diagnosis , Child, Preschool , Electric Impedance , Female , Humans , Hydrogen-Ion Concentration , Infant , MaleABSTRACT
In this study, γ-glutamyl transpeptidase (GGT) and l-cysteine sulphoxide lyase (C-S lyase) were purified from the fruiting body of Lentinula edodes in three steps and then characterised. We found that GGT together with C-S lyase caused the generation of endogenous formaldehyde in L. edodes. GGT was composed of a large subunit of 41 kDa and a small subunit of 25 kDa, and C-S lyase was composed of two identical subunits of 46 kDa, as determined by SDS-PAGE. GGT was stable at pH 8.0-10.0 with an optimum pH of 8.8, and was stable at 20-50°C with an optimum activity at 37°C. C-S lyase was stable at pH 8.0-9.0 with an optimum pH of 8.5, and was stable at 20-60°C with an optimum activity at 40°C. The present work supports the study of the mechanism of endogenous formaldehyde in L. edodes.
