Isolation and Characterization of a Molybdenum Reducing Enzyme in Enterobacter cloacae Strain 48
Molybdenum reducing enzyme was isolated from Enterobacter cloacae Strain 48 by ammonium sulphate fractionation, DE-cellulose ion-exchange chromatography and Sephacryl S-200 gel filtration. SDS-PAGE of the concentrated Sephacryl S-200 gel filtration eluates revealed the presence of 3 protein subu...
| Main Authors: | , , , , , |
|---|---|
| Format: | Article |
| Language: | English English |
| Published: |
Universiti Putra Malaysia Press
2003
|
| Online Access: | http://psasir.upm.edu.my/id/eprint/3778/1/Isolation_and_Characterization_of_a_Molybdenum_Reducing.pdf |
| _version_ | 1796966618804256768 |
|---|---|
| author | Abd. Shukor, Mohd. Yunus Lee, C. H. Omar, Ismail Karim, M. I. A. Syed, Mohd Arif Shamaan, Nor Aripin |
| author_facet | Abd. Shukor, Mohd. Yunus Lee, C. H. Omar, Ismail Karim, M. I. A. Syed, Mohd Arif Shamaan, Nor Aripin |
| author_sort | Abd. Shukor, Mohd. Yunus |
| collection | UPM |
| description | Molybdenum reducing enzyme was isolated from Enterobacter cloacae Strain 48
by ammonium sulphate fractionation, DE-cellulose ion-exchange
chromatography and Sephacryl S-200 gel filtration. SDS-PAGE of the
concentrated Sephacryl S-200 gel filtration eluates revealed the presence of 3
protein subunits of molecular weight 80, 90 and 100 kDa. The active
concentrated fraction from the Sephacryl S-200 gel filtration step was then
characterized for molybdenum reducing activity with 12-molybdophosphate
(12-MoP) as a substrate. The optimum pH and temperature of the reaction was
5.0 and 28-33°C, respectively. ADH was a better reducing agent in the
reaction than NADPH; the double reciprocal plot of activity against ADH and
NADPH revealed apparent Km and V""", values of 1.65 mM, 6.28 nmole
molybdenum blue produced/min/mg and 2.13 mM and 4.10 nmole
molybdenum blue produced/min/mg, respectively. The double reciprocal plot
of activity against 12-MoP and 20-molybdodiphosphate revealed apparent K
m values of 0.3 mM and 0.4 mM, respectively. The apparent Vmax values are similar
for both substrates at 6 nmole molybdenum blue produced/min. The assay
method for molybdenum reducing activity using 12-MoP was found to be easier
and more rapid than the present method of using molybdate as a substrate |
| first_indexed | 2024-03-06T07:02:07Z |
| format | Article |
| id | upm.eprints-3778 |
| institution | Universiti Putra Malaysia |
| language | English English |
| last_indexed | 2024-03-06T07:02:07Z |
| publishDate | 2003 |
| publisher | Universiti Putra Malaysia Press |
| record_format | dspace |
| spelling | upm.eprints-37782013-05-27T07:11:09Z http://psasir.upm.edu.my/id/eprint/3778/ Isolation and Characterization of a Molybdenum Reducing Enzyme in Enterobacter cloacae Strain 48 Abd. Shukor, Mohd. Yunus Lee, C. H. Omar, Ismail Karim, M. I. A. Syed, Mohd Arif Shamaan, Nor Aripin Molybdenum reducing enzyme was isolated from Enterobacter cloacae Strain 48 by ammonium sulphate fractionation, DE-cellulose ion-exchange chromatography and Sephacryl S-200 gel filtration. SDS-PAGE of the concentrated Sephacryl S-200 gel filtration eluates revealed the presence of 3 protein subunits of molecular weight 80, 90 and 100 kDa. The active concentrated fraction from the Sephacryl S-200 gel filtration step was then characterized for molybdenum reducing activity with 12-molybdophosphate (12-MoP) as a substrate. The optimum pH and temperature of the reaction was 5.0 and 28-33°C, respectively. ADH was a better reducing agent in the reaction than NADPH; the double reciprocal plot of activity against ADH and NADPH revealed apparent Km and V""", values of 1.65 mM, 6.28 nmole molybdenum blue produced/min/mg and 2.13 mM and 4.10 nmole molybdenum blue produced/min/mg, respectively. The double reciprocal plot of activity against 12-MoP and 20-molybdodiphosphate revealed apparent K m values of 0.3 mM and 0.4 mM, respectively. The apparent Vmax values are similar for both substrates at 6 nmole molybdenum blue produced/min. The assay method for molybdenum reducing activity using 12-MoP was found to be easier and more rapid than the present method of using molybdate as a substrate Universiti Putra Malaysia Press 2003 Article PeerReviewed application/pdf en http://psasir.upm.edu.my/id/eprint/3778/1/Isolation_and_Characterization_of_a_Molybdenum_Reducing.pdf Abd. Shukor, Mohd. Yunus and Lee, C. H. and Omar, Ismail and Karim, M. I. A. and Syed, Mohd Arif and Shamaan, Nor Aripin (2003) Isolation and Characterization of a Molybdenum Reducing Enzyme in Enterobacter cloacae Strain 48. Pertanika Journal of Science & Technology, 11 (2). pp. 261-272. ISSN 0128-7680 English |
| spellingShingle | Abd. Shukor, Mohd. Yunus Lee, C. H. Omar, Ismail Karim, M. I. A. Syed, Mohd Arif Shamaan, Nor Aripin Isolation and Characterization of a Molybdenum Reducing Enzyme in Enterobacter cloacae Strain 48 |
| title | Isolation and Characterization of a Molybdenum Reducing
Enzyme in Enterobacter cloacae Strain 48 |
| title_full | Isolation and Characterization of a Molybdenum Reducing
Enzyme in Enterobacter cloacae Strain 48 |
| title_fullStr | Isolation and Characterization of a Molybdenum Reducing
Enzyme in Enterobacter cloacae Strain 48 |
| title_full_unstemmed | Isolation and Characterization of a Molybdenum Reducing
Enzyme in Enterobacter cloacae Strain 48 |
| title_short | Isolation and Characterization of a Molybdenum Reducing
Enzyme in Enterobacter cloacae Strain 48 |
| title_sort | isolation and characterization of a molybdenum reducing enzyme in enterobacter cloacae strain 48 |
| url | http://psasir.upm.edu.my/id/eprint/3778/1/Isolation_and_Characterization_of_a_Molybdenum_Reducing.pdf |
| work_keys_str_mv | AT abdshukormohdyunus isolationandcharacterizationofamolybdenumreducingenzymeinenterobactercloacaestrain48 AT leech isolationandcharacterizationofamolybdenumreducingenzymeinenterobactercloacaestrain48 AT omarismail isolationandcharacterizationofamolybdenumreducingenzymeinenterobactercloacaestrain48 AT karimmia isolationandcharacterizationofamolybdenumreducingenzymeinenterobactercloacaestrain48 AT syedmohdarif isolationandcharacterizationofamolybdenumreducingenzymeinenterobactercloacaestrain48 AT shamaannoraripin isolationandcharacterizationofamolybdenumreducingenzymeinenterobactercloacaestrain48 |