Human Liver High Molecular Weight Zinc-Dependent Acid p-Nitrophenylphosphatase. Purification and Properties.
Bibliographic Information
- Other Title
-
- Human Liver High Molecular Weight Zinc-
Search this article
Abstract
Human liver contains high molecular weight-type Zn2+-dependent acid p-nitrophenylphosphatase (HMW-ZnAP). The enzyme was purified 1000-fold by a new procedure, including preparative isoelectrofocusing. The HMW-ZnAP was homogeneous in non-denaturing disk-gel electrophoresis with an MW of about 93 kDa determined by Sephadex G-100 chromatography. A single polypeptide chain of 43 KDa was detected on sodium dodecyl sulphate polyacrylamide gel electrophoresis (SDS-PAGE), suggesting a homodimeric structure. The isoelectric point (pl) was 7.2-7.4. Human liver HMW-ZnAP requires Zn2+-ions for activity; other divalent cations are ineffective or act as inhibitors. It dephosphorylated p-nitrophenylphosphate (pNPP) (Km=0.24 mM), o-carboxyl phenylphosphate (oCPP) (Km=0.92 mM) and phenylphophate (PhP) (Km=1.42 mM). Other substrates including [32P]-labelled casein or phosvitin, adenyl nucleotides and myo-inositol-1-phosphate, were not dephosphorylated. Human liver HMW-ZnAP obeys Michaelis-Menten kinetics with pNPP as substrate; the enzyme was competitively inhibited by inorganic phosphate (Ki=0.55 mM), and by oCPP (Ki=0.65 mM) and PhP (Ki=1.16 mM). Adenosine monophosphate (AMP), adenosine diphosphate (ADP) and ATP displayed mixed-type inhibition. The enzyme was also inhibited by some modifiers such as EDTA, oxalate, p-chloromercurybenzoate, tartrate, imidazole, cyanide, cysteine, histidine and diethylpyrocarbonate, but not by fluoride or okadaic acid. Human liver HMW-ZnAP is sensitive to temperatures higher than 40°C. The pH-dependence of the steady-state kinetic parameters indicates the existence of an essential ionizable group with a pKa of 7.25-7.50, similar to that of histidine. However, diethylpyrocarbonate inactivation experiments suggest that other amino acid residues may also be involved in enzyme catalysis.
Journal
-
- Biological and Pharmaceutical Bulletin
-
Biological and Pharmaceutical Bulletin 20 (12), 1235-1239, 1997
The Pharmaceutical Society of Japan
- Tweet
Keywords
Details 詳細情報について
-
- CRID
- 1390282679599943680
-
- NII Article ID
- 110003638988
-
- NII Book ID
- AA10885497
-
- ISSN
- 13475215
- 09186158
-
- NDL BIB ID
- 4371357
-
- PubMed
- 9448095
-
- Text Lang
- en
-
- Data Source
-
- JaLC
- NDL
- Crossref
- PubMed
- CiNii Articles
-
- Abstract License Flag
- Disallowed