Malaysian Applied Biology Journal

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36-2-01

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Malays. Appl. Biol. (2007) 36(2): 1–5

A RAPID SCREENING METHOD FOR CGTASE-PRODUCING BACTERIA USING DIFFERENT STARCHES AS CARBON SOURCE

SURAINI ABD-AZIZ*1, SAUVAPHAP AI NOI1, OSMAN HASSAN2, MOHAMMED ISMAIL ABDUL KARIM3, NORJAHAN BANU ALITHEEN1, KAMARULZAMAN KAMARUDDIN4 and ROSLI MD. ILLIAS5

1 Faculty of Biotechnology and Biomolecular Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia
2 Faculty of Science and Technology, School of Chemical Sciences and Food Technology, Universiti Kebangsaan Malaysia, 43600 Bangi, Selangor, Malaysia
3 Department of Biotechnology Engineering, Faculty of Engineering, International Islamic University of Malaysia, Jalan Gombak, 53100 Kuala Lumpur, Malaysia
4Bioprocess and Chemical Technology Centre, SIRIM Berhad, P.O. Box 7035, Section 2, 40911 Shah Alam, Selangor, Malaysia
5 Faculty of Chemical Engineering and Natural Resources Engineering, Universiti Teknologi Malaysia, 81310 Skudai, Johor, Malaysia

ABSTRACT

Cyclodextrin glycosyltransferases (CGTases) are extracellular bacterial enzymes that generate cyclodextrin from starch. Screening, isolation and characterisation of CGTases are extensively carried out due to its importance in industrial biotechnology. Conventionally, identification of CGTase-producing bacteria involves the use of solid media containing phenolphthalein-methyl orange as indicators by colour changes. The formation of CGTase does not require a specific inducer as the presence of starch is essentially sufficient. In the present study, modification of the conventional method was developed by substituting soluble starch with five other types of starches, namely, corn, rice, tapioca, glutinous starch and sago. The improved method enables simultaneous isolation and screening for CGTase-producing bacteria and identification of the most suitable carbon source (starch) for enzyme production. The diameter of the clearing zone formation around the bacterial colony in the starch-containing medium is used to gauger the hydrolytic efficiency of the bacteria. Out of the 250 soil bacterial samples screened, strain MK 6 was identified as the most prolific CGTase producer. This isolated strain was capable of hydrolysing sago starch best (and least in glutinous starch). The modified method using simple substrate substitution and the phenolphthalein colour reaction studied here was thus found to be useful for the rapid and qualitative method for preliminary screening of CGTase-producing bacteria.


ABSTRAK

Siklodekstrin glikosiltransferase (CGTases) adalah enzim bakteria ekstraselular yang bertanggungjawab dalam penghasilan siklodekstrin daripada kanji. Penyaringan, pemencilan dan pencirian yang ekstensif dijalankan ke atas CGTase berdasarkan kepentingannya dalam bidang bioteknologi industri. Kebiasaanya, penentuan bakteria penghasil CGTase melibatkan penggunaan media pepejal yang mengandungi phenolphthalein-methyl orange sebagai penanda bagi perubahan warna yang berlaku. Pembentukan CGTase tidak memerlukan pengalak spesifik kerana kehadiran kanji di dalam sistem adalah mencukupi. Di dalam kajian ini, pengubahsuaian ke atas kaedah sedia ada telah dilakukan melalui penggantian kanji terlarut dengan lima jenis kanji terpilih seperti kanji jagung, beras, keledek, pulut dan sagu. Pengubahsuaian kaedah ini membolehkan penyaringan dan pemencilan bakteria penghasil CGTase dapat dilakukan dengan mudah dan penentuan sumber karbon yang paling sesuai dapat dikenalpasti bagi penghasilan enzim. Pembentukan kawasan zon cerah di keliling koloni bakteria pada media yang mengandungi kanji dapat digunakan sebagai pengukur keberkesanan hidrolisis oleh bakteria. Daripada 250 sampel bakteria yang disaring, strain MK 6 telah dikenalpasti dan dipilih sebagai penghasil CGTase yang berpotensi. Strain yang dipencilkan ini berupaya menghidrolisis kanji sagu dengan baik dan diakhiri dengan kanji pulut. Kaedah terubahsuai ini menggunakan substrat ringkas dan tindakbalas perubahan warna fenolftalein didapati berguna untuk kaedah yang cepat dan kualitatif dalam proses penyaringan awal bakteria penghasil CGTase.

Key words: Rapid screening method, CGTase, Bacteria, Carbon, Starch

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