Native Malate Dehydrogenase from Microorganism

Products

Native Malate Dehydrogenase from Microorganism

Product Specifications

Cat#	NATE-1016
Descriptions	High-purity Malate Dehydrogenase mediates malate and oxaloacetate redox cycles as a versatile auxiliary enzyme for multiple clinical metabolite detection assays. It exhibits high reaction conversion efficiency and low impurity interference, widely matched to diagnostic reagent formulas for amino acid, energy metabolism and organic acid quantitative analysis in clinical laboratories.
Applications	This product can be used in clinical diagnosis for enzymatic quantification of L-malate and activity detection of aspartate aminotransferase (AST).
Synonyms	(S)-Malate: NAD⁺ oxidoreductase
EC Number	EC 1.1.1.37
Catalysis	L-Malate + NAD⁺ ⇆ Oxaloacetate + NADH + H⁺
Activity	≥ 100 U/mg (Malate dehydrogenase activity)
Contaminant	≤ 4 × 10⁻⁴% (NADH oxidase activity); ≤ 3 × 10⁻³% (α-KG dehydrogenase activity); ≤ 3 × 10⁻³% (GOT activity)
Appearance	White powder, lyophilized
Additive	Dextran
Molecular Weight	35,000 (SDS-PAGE)
Isoelectric Point	4.8
Km	5.6 × 10⁻⁵ M (Oxaloacetate); 6.6 × 10⁻⁵ M (NADH)
Optimum pH	7.8
pH Stability	8.0-9.0 (70°C, 30 min)
Optimum Temperature	70°C
Thermal Stability	up to 80°C (pH 8.5, 30 min)
Inhibitors	Zn²⁺
Unit Definition	1 unit is defined as the enzyme quantity which oxidizes 1 μ mole of NADH per minute under the conditions described below.
Storage	18 months from the date of analysis when stored at 5°C or below in a dry place under sealed conditions.

General Properties

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Assay Principle

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Activity Assay

Unit Definition

1 unit is defined as the enzyme quantity which oxidizes 1 μ mole of NADH per minute under the conditions described below.

Reagents

A. 0.1 M Tris-HCl buffer (pH 7.8)
B. 0.01 M Phosphate buffer (KH2PO4-NaOH, pH 7.0)
C. Triton X-100 solution (50 mg/mL)
D. 0.01 M Phosphate buffer containing 0.1% Triton X-100 (KH2PO4-NaOH, pH 7.0): Dilute 20 mL of Triton X-100 solution (C) with approx. 800 mL of 0.01M Phosphate buffer (B). Fill up to 1,000 mL with 0.01M Phosphate buffer (B).
E. NADH solution: Weigh 9 mg of NADH and dissolve in 0.1 M Tris-HCl buffer (A). Fill up to 50 mL with 0.1 M Tris HCl Buffer (A). (Can be used for 5 days if kept refrigerated)
F. Substrate solution: Weigh 11 mg of oxaloacetic acid and dissolve in 0.1 M Tris-HCl buffer (A). Fill up to 50 mL with 0.1 M Tris-HCl buffer (A). (Make a fresh solution for each use.)
G. Enzyme solution: Weigh out Malate Dehydrogenase and dissolve in chilled 0.01M Phosphate Buffer containing 0.1% Triton X100(D). Enzyme solution should be prepared so that the value of ΔOD/minute becomes in the range of 0.025 ± 0.010.

Procedure

Pipette 2.0 mL of NADH solution (E) and 0.90 mL of Substrate solution (F) respectively into a quartz cell (d=10 mm) and keep at 25 ± 0.5℃ for 5 minutes. Then, pipette 0.10 mL of Enzyme solution (G) into the quartz cell and mix well immediately. Keep the reaction mixture at 25 ± 0.5℃. Exactly at 2 minutes and 5 minutes after the addition of Enzyme solution (G), measure the absorbances of the reaction mixture at 340 nm (A2 and A5). As a blank, pipette 0.01M Phosphate buffer (D) into another quartz cell (d=10 mm) instead of the Enzyme solution (G) and follow the same procedure described above (Ab2 and Ab5).

Calculation

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Safe Handling Avoid inhalation; in the event of direct skin or eye contact, immediately flush the affected area with abundant water, and consult the SDS for further information.