Occurrence of myo-inositol-1-phosphate phosphatase in pteridophytes: characteristics of the enzyme from the reproductive pinnules of Dryopteris filix-mas (L.) Schott

Abstract

Evident myo-inositol-1-phosphate phosphatase (MIPP) activity has been detected both in the vegetative as well as in the spore-bearing organs of some selected pteridophytes having wide phylogenetic diversity. The basic characterization of this enzyme was carried out using the cosmopolitan fern Dryopteris filix-mas. The enzyme was partially purified from the cytosol fraction obtained from the reproductive pinnules of the plant to about 41-fold over the initial homogenate following low-speed centrifugation, streptomycin sulfate precipitation, 25-70% ammonium sulfate fractionation, CM Sephadex C-50 chromatography and finally gel-filtration on Ultrogel AcA 34. The apparent molecular weight of the native MIPP was estimated to be 94 kDa. The enzyme activity increased linearly with respect to protein concentration to about 150 µg and with respect to time up to 75 min. The temperature optimum was found at 40ºC. However, the enzyme showed good activity over the temperature range of 30-50ºC. This enzyme used D/L-myo-inositol-1-phosphate as its principal substrate (95-100%), however, about 16% activity was recorded when D-myo-inositol-3-phosphate substituted as substrate. Furthermore, weak (3%) activity of this MIPP was observed with 2-glycerophosphate as substrate. The apparent K m for pteridophytic MIPP was 0.083 mM. The enzyme was functional in a narrow pH range of 7.5 to 8.5. The activity of this MIPP enzyme was remarkably inhibited by the presence of a monovalent cation, lithium, and even moderately so at a low concentration such as 1 mM. On the other hand, magnesium, a divalent cation, enhanced activity at least up to 10 mM. Calcium diminished MIPP activity at concentrations over 4 mM. Key words: Dryopteris filix-mas, myo-inositol-1-phosphate phosphatase, pteridophytes, reproductive pinnulesOcorrência da fosfatase do mio-inositol-1-fosfato em pteridófitas: características da enzima a partir de pínulas reprodutivas de Dryopteris filix-mas (L.) Schott: Tem-se detectado atividade da fosfatase do mio-inositol-1-fosfato (FMIF) tanto em órgãos vegetativos como em estruturas esporulantes de algumas pteridófitas com ampla diversidade filogenética. Neste estudo, procedeu-se à caracterização básica dessa enzima utilizando-se da pteridófita cosmopolita Dryopteris filix-mas. Após centrifugação em baixa velocidade, precipitação com sulfato de estreptomicina, fracionamento com sulfato de amônio (25-70%), cromatografia em CM Sephadex C-50 e, finalmente, filtração gélica em Ultrogel AcA 34, conseguiu-se uma purificação parcial da enzima (a partir da fração citossólica obtida de pínulas reprodutivas da planta) de cerca de 41 vezes em relação ao homogenato inicial. O peso molecular aparente da FMIF nativa foi estimado em 94 kDa. A atividade da enzima aumentou linearmente com relação ao conteúdo de proteína (cerca de 150 µg) e com relação ao tempo (até 75 min). A temperatura ótima foi de 40ºC. Entretanto, a enzima exibiu atividade razoável na faixa de temperatura entre 30 e 50ºC. O D/L-mio-inositol-1-fosfato foi o principal substrato (9...

Biosynthesis of Myo-Inositol in Chloroplasts of Salinity-Stressed Marine Macro Alga Ulva lactuca

“…Perusal of available literature clearly reveals that the molecular weight of native plastidial MIPS has been found to vary between 155 kDa (Diplopterygium glaucum) (Chettri et al, 2006b) and 180 kDa (Entamoeba histolytica) (Lohia et al, 1999). The molecular weight of native cytosolic MIPP from a common fern Dryopteris filix-mas has been reported to be around 94 kDa (Banerjee et al, 2007), but no specific information is available regarding the molecular weight of its plastidial iso-form in that fern genus. In the present communication, however, an attempt was made to fulfil this lacunae existing in previous reports.…”

“…L-myo-inositol-1-phosphate synthase (MIPS) carries out the irreversible conversion of D-Glucose-6-phosphate (G-6-P) to L-myo-inositol-1-phosphate (I-1-P), which is subsequently dephosphorylated by inositol monophosphatase (MIPP) to free myo-inositol (MI) (eisenBerg, 1967).Together these two enzymes interact with each other in inositol biosynthesis metabolon. The MIPS and MIPP reactions have been reported from several pro-as well as eukaryotic sources covering a wide phylogenetic diversity (ei-senBerg, 1967;Maeda & eisenBerg, 1980;Loewus & Loewus, 1983;adhikari & MajuMder, 1983;guMBer et al, 1984;Lohia et al, 1999;Chen et al, 2000;MajuMder & Biswas 2006;Chhetri et al, 2006a, b;Patra et al, 2007;Banerjee et al, 2007;Basak et al, 2012). In this context, it is noteworthy to mention that although few studies on MIPS have been performed from various living systems inclusive of plants; however, studies concerning MIPP, specifically in plants, are surprisingly very limited.…”

“…In this context, it is noteworthy to mention that although few studies on MIPS have been performed from various living systems inclusive of plants; however, studies concerning MIPP, specifically in plants, are surprisingly very limited. Although the inositol biosynthetic pathway has been found to be operative in vascular cryptogams and flowering plants (Loewus & Loewus, 1983;guMBer et al, 1984;adhikari et al, 1987;Chhetri et al, 2006a, b;Banerjee et al, 2007;Basak et al, 2012), specific information regarding its functionality in non-vascular cryptogams is a neglected area of research. Taking these facts into consideration, attempts were made in the present investigation on purification and characterization of chloroplast localized MIPS and MIPP from a Chlorophycean marine macroalga Ulva lactuca Linn.…”

Pengaruh Myoinositol dan Arang Aktif terhadap Pertumbuhan Planlet Anggrek Dendrobium dalam Kultur In Vitro

“…Peningkatan pertumbuhan panjang dan lebar daun pada perlakuan myoinositol 50 mg/l disebabkan karena adanya percepatan pembelahan sel dan mendorong proses diferensiasi. Pembelahan sel membutuhkan energi tinggi yang diperoleh dari myoinositol karena inositol ialah golongan karbohidrat (Barnejee et al 2007). …”

“…Myoinositol merupakan senyawa karbohidrat meskipun bukan merupakan gula pada umumnya (Barnejee et al 2007). Karbohidrat selain sebagai bahan baku yang menghasilkan energi untuk proses respirasi, juga sebagai bahan pembentuk sel-sel baru, dalam konsentrasi yang tepat dapat merangsang perakaran.…”