Membrane-bound class III peroxidases: identification, biochemical properties and sequence analysis of isoenzymes purified from maize (Zea mays L.) roots.
The occurrence of three plasma membrane-bound class III peroxidases has been demonstrated for maize (Zea mays L.) roots [Mika and Lüthje (2003) Plant Physiol. 132:1489-1498]. In the present work a novel PM-bound peroxidase (pmPOX3) was partially purified. The experimental molecular mass of the heme protein was 38 kDa after size exclusion, and 57 kDa in non-reducing SDS-PAGE stained with the peroxidase substrates tetramethylbenzidine and H(2)O(2). The glycosylation of pmPOX1, pmPOX2b and pmPOX3 was shown by different approaches. The full length sequences of pmPOX1, pmPOX2b and pmPOX3 were identified by ESI-MS/MS and MALDI-TOF MS analysis in combination with in silico and in vivo cloning. Thus, we report the first sequence analysis of membrane-bound class III peroxidases. A partial gene analysis revealed two or three introns. Experimental and theoretical isoelectric points and molecular masses were compared. Targeting signals, the putative protein structures and the localization of the active center of the enzymes on the outside of the plasma membrane were deduced of the amino acid sequences. In contrast to other class III peroxidases, pmPOX1 seems to have a dimeric structure. Predictions of hydrophobic domains in comparison with solubilization experiments suggest an N-terminal transmembrane domain for the isoenzymes.
Mika
Angela
A
University of Hamburg, Biocenter Klein Flottbek and Botanical Garden, Plant Physiology, Ohnhorststrasse 18, D-22609 Hamburg, Germany.
Buck
Friedrich
F
Lüthje
Sabine
S
D000595:Amino Acid Sequence; D001483:Base Sequence; D002462:Cell Membrane / Q000201:enzymology; D002850:Chromatography, Gel; D006031:Glycosylation; D007526:Isoelectric Point; D007527:Isoenzymes / Q000737:chemistry / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism; D008958:Models, Molecular; D008969:Molecular Sequence Data; D008970:Molecular Weight; D010544:Peroxidases / Q000737:chemistry* / Q000235:genetics / Q000302:isolation & purification / Q000378:metabolism*; D018517:Plant Roots / Q000201:enzymology* / Q000235:genetics; D011485:Protein Binding; D017434:Protein Structure, Tertiary; D017421:Sequence Analysis; D017386:Sequence Homology, Amino Acid; D040681:Structural Homology, Protein; D053719:Tandem Mass Spectrometry; D003313:Zea mays / Q000201:enzymology* / Q000235:genetics
D007527:Isoenzymes; D010544:Peroxidases