Purification and biochemical characterization of two major thermophilic xylanase isoforms (T 70 and T 90) from xerophytic Opuntia vulgaris plant spp

Research output: Journal Publications and Reviews (RGC: 21, 22, 62)21_Publication in refereed journalpeer-review

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Author(s)

  • Jeyaraman Vikramathithan
  • Sambandam Ravikumar
  • Pandurangan Muthuraman
  • Gali Nirmalkumar
  • Sivalingam Shayamala
  • And 1 others
  • Kotteazeth Srikumar

Related Research Unit(s)

Detail(s)

Original languageEnglish
Pages (from-to)1373-1383
Journal / PublicationCellulose
Volume19
Issue number4
Online published5 Apr 2012
Publication statusPublished - Aug 2012

Abstract

Thermostable xylanase isoforms T 70 and T 90 were purified and characterized from the xerophytic Opuntia vulgaris plant species. The enzyme was purified to homogeneity employing three consecutive steps. The purified T 70 and T 90 isoforms yielded a final specific activity 134. 0 and 150. 8 U mg -1 protein, respectively. The molecular mass of these isoforms was determined to be 27 kDa. The optimum pH for the T 70 and T 90 xylanase isoforms was 5. 0 and the temperature for optimal activity was 70 and 90 °C, respectively. The Km value of T 70 and T 90 enzyme isoforms was 3. 49, 2. 1 mg ml -1, respectively when oat spelt xylan was used as a substrate. The T 70 had a Vmax of 10. 4 μmol min -1 mg -1, and T 90 had a Vmax of 8. 9 μmol min -1 mg -1, respectively. In the presence of 10 mM Co 2+, and Mn 2+ the activity of T 70 and T 90 isoforms increased, where as 90 % inhibition was noted with of the use 10 mM Hg 2+, Cd 2+, Cu 2+, Zn 2+ while partial inhibition was observed in the presence of Fe 3+, Ni 2+, Ca 2+and Mg 2+. The T 70 and T 90 isoforms retained nearly 50 % activity in the presence of 2. 0 M urea, while use of 40 mM SDS lowered the activity nearly 38-41 %. The substrate specificity of both T 70 and T 90 isoforms showed maximum activity for oat spelt xylan. Western blot, immunodiffusion, and in vitro inhibition assays confirmed reactivity of the T 90 isoform with polyclonal anti-T 90 antibody raised in rabbit, as well as cross-reactivity of the antibody with the T 70 xylanase isoform.

Research Area(s)

  • Metal ions, Oat spelt, Opuntia vulgaris, Protein denaturants, Thermophilic, Xylanase

Citation Format(s)

Purification and biochemical characterization of two major thermophilic xylanase isoforms (T 70 and T 90) from xerophytic Opuntia vulgaris plant spp. / Vikramathithan, Jeyaraman; Ravikumar, Sambandam; Muthuraman, Pandurangan; Nirmalkumar, Gali; Shayamala, Sivalingam; Srikumar, Kotteazeth.

In: Cellulose, Vol. 19, No. 4, 08.2012, p. 1373-1383.

Research output: Journal Publications and Reviews (RGC: 21, 22, 62)21_Publication in refereed journalpeer-review