Identification and characterization of L-lysine decarboxylase from Huperzia serrata and its role in the metabolic pathway of lycopodium alkaloid.

Lysine decarboxylation is the first biosynthetic step of Huperzine A (HupA). Six cDNAs encoding lysine decarboxylases (LDCs) were cloned from Huperzia serrata by degenerate PCR and rapid amplification of cDNA ends (RACE). One HsLDC isoform was functionally characterized as lysine decarboxylase. The HsLDC exhibited greatest catalytic efficiency (kcat/Km, 2.11 s-1 mM-1) toward L-lysine in vitro among all reported plant-LDCs. Moreover, transient expression of the HsLDC in tobacco leaves specifically increased cadaverine content from zero to 0.75 mg per gram of dry mass. Additionally, a convenient and reliable method used to detect the two catalytic products was developed. With the novel method, the enzymatic products of HsLDC and HsCAO, namely cadaverine and 5-aminopentanal, respectively, were detected simultaneously both in assay with purified enzymes and in transgenic tobacco leaves. This work not only provides direct evidence of the first two-step in biosynthetic pathway of HupA in Huperzia serrata and paves the way for further elucidation of the pathway, but also enables engineering heterologous production of HupA.

Hyperoxaluria is reduced and nephrocalcinosis prevented with an oxalate-degrading enzyme in mice with hyperoxaluria.

BACKGROUND/AIMS: Hyperoxaluria is a major risk factor for recurrent urolithiasis and nephrocalcinosis. We tested an oral therapy with a crystalline, cross-linked formulation of oxalate-decarboxylase (OxDc-CLEC) on the reduction of urinary oxalate and decrease in the severity of kidney injury in two models: AGT1 knockout mice (AGT1KO) in which hyperoxaluria is the result of an Agxt gene deficiency, and in AGT1KO mice challenged with ethylene glycol (EG). METHODS: Four different doses of OxDc-CLEC mixed with the food, or placebo were given to AGT1KO mice (200 mg/day, n = 7) for 16 days and to EG-AGT1KO mice (5, 25, and 80 mg, n = 11) for 32 days. RESULTS: Oral therapy with 200 mg OxDc-CLEC reduced both urinary (44%) and fecal oxalate (72%) in AGT1KO mice when compared to controls. Similarly, in EG-AGT1KO mice, each of the three doses of OxDc-CLEC produced a 30-50% reduction in hyperoxaluria. A sustained urinary oxalate reduction of 40% or more in the 80 mg group led to 100% animal survival and complete prevention of nephrocalcinosis and urolithiasis. CONCLUSION: These data suggest that oral therapy with OxDc-CLEC may reduce hyperoxaluria, prevent calcium oxalate nephrocalcinosis and urolithiasis, and can represent a realistic option for the treatment of human hyperoxaluria, independent of cause.

Efecto de la cocarboxilasa no degradable en la acidosis metabólica del recién nacido / Metabolic acidosis in the neonate: Efficacy of non degradable cocarboxylase

Antecedentes: La cocarboxilasa no degradable (CND), es una coenzima importante para la síntesis de la deshidrogenasa pirúvica, enzima que regula el catabolismo del piruvato y posiblemente la clave para la recuperación de la acidosis metabólica. Objetivo: Evaluar la eficacia de la cocarboxilasa no degradable para manejar la acidosis metabólica del recién nacido. Material y métodos: Se realizó un estudio doble ciego controlado, seleccionado aleatoriamente 30 neonatos durante un año, con diagnóstico gasométrico de acidosis metabólica (pH de 7.30 o menos, bicarbonato de 18 o menos, y exceso de base de -9 o mayor). Se administró solución isotónica de cloruro de sodio (grupo I) 1ml/kg, bicarbonato de sodio (grupo II) 2mEq/kg y CND (grupo III) a razón de 40 mg/kg. El volumen en todos los casos fue de 1 mL/kg. La primera dosis se administró en cuanto se realizó el diagnóstico y las dos siguientes con espacio de 8 horas. Se determinó el efecto de las sustancias en el pH y la corrección de la acidosis metabólica. Resultados: Los resultados mostraron un discreto aumento del pH con la CND sin diferencias para ninguna de las sustancias sobre la acidosis metabólica

Induction of aromatic-L-amino acid decarboxylase by decarboxylase inhibitors in idiopathic parkinsonism.

We evaluated the effect of administration of L-dopa, alone or in combination with a peripheral decarboxylase inhibitor, on plasma levels of aromatic-L-amino acid decarboxylase (ALAAD). After single-dose administration of L-dopa plus benserazide (Madopar) in healthy subjects and in chronically treated patients with parkinsonism, plasma ALAAD followed for 2 to 3 hours fell, but returned to predosing levels within 90 minutes. Four groups of patients with idiopathic parkinsonism were studied during chronic treatment: Group I, no L-dopa treatment (n = 31); Group II, L-dopa alone (n = 15); Group III, L-dopa plus benserazide (n = 28); and Group IV, L-dopa plus carbidopa (Sinemet, n = 30). Plasma ALAAD 2 hours after dosing was normal in Groups I and II. ALAAD was increased threefold in Groups III and IV, suggesting induction of ALAAD by the coadministration of a peripheral decarboxylase inhibitor. In a study of 3 patients in whom L-dopa/benserazide was started, plasma ALAAD rose gradually over 3 to 4 weeks. Further detailed pharmacokinetic studies of L-dopa, dopamine, and ALAAD in plasma and cerebrospinal fluid are required to determine if the apparent ALAAD induction by a peripheral decarboxylase inhibitor may be related to the loss of clinical efficacy of combination therapy in some patients and how it is related to end-of-dose deterioration and on-off phenomena.