Molecular Cloning of Phytase Gene from ASUIA279 and Its Expression in Pichia pastoris System

Phytases catalyze the hydrolysis of phytate (myo-inositol hexakisphosphate), one of the major storage form of phosphate in plants, with subsequent release of myo-inositol, phosphate and phytate-bound minerals. Non-ruminant animals such as chicken, swine and fish can't use the organic phosphorus and minerals from their diet because there is no phytase activity in their digestive tract. Phytate degrading enzyme is added to the animal feed diet to improve phosphorus availability from the dietary phytate and at the same time this lessen the phosphate pollution level in areas of intensive animal production as the phosphate would not be excreted out to the environment. ASUIA279, a bacterial strain isolated from Malaysian soil has potentially shown good phytase activity. In the present work, the gene encoding for phytase has been amplified from the plasmid DNA of recombinant ASUIA279(5) obtained from a previous study (unpublished data) by a polymerase chain reaction (PCR) methodology. The amplified phytase gene was extracted, purified then cloned into the pPICZA plasmid and transformed into Pichia pastorisX-33 strain for enzyme production. ABSTRAK: Fitase pemangkinan hidrolisis fitat (myo-inositol hexakisphosphate), merupakan salah satu cara penyimpanan utama fosfat dalam tumbuhan, dengan pelepasan berturut myo-inositol , fosfat dan galian terikat fitat. Haiwan bukan ruminan seperti ayam, khinzir dan ikan tidak dapat memanfaatkan fosforus organik serta galian yang diperolehi daripada makanan kerana tidak mempunyai aktiviti fitase di dalam saluran pencernaan mereka. Enzim pengecilan fitase dicampurkan ke dalam pemakanan haiwan untuk mempertingkatkan keperolehan fosforus dari fitat diet. Pada masa yang sama ia dapat mengurangkan tahap pencemaran fosfat di kawasan yang terdapat penternakan haiwan secara intensif agar fosfat tidak dikumuhkan ke persekitaran. ASUIA279, satu strain bakteria yang diasingkan daripada tanih di Malaysiamenunjukkan aktiviti fitase yang memberansangkan. Buat masa kini, pengekodan gen fitase telah diperkuatkan dengan plasmid DNA dari ASUIA279(5), rekombinan yang diperolehi daripada kajian terdahulu (data tak diterbitkan) berdasarkan kaedah tindak balas berantai polimerase (polymerase chain reaction (PCR)). Gen fitase yang diperkuatkan, ditulenkan dan kemudian diklonkan menjadi plasmid pPICZαA dan berubah menjadi strain Pichia pastorisX-33 untuk penghasilan enzim. KEYWORDS:  Phytase, animal feed, pollution, polymerase chain reaction (PCR), Pichia pastori

Untargeted metabolomics analysis reveals key pathways responsible for the synergistic killing of colistin and doripenem combination against Acinetobacter baumannii

Combination therapy is deployed for the treatment of multidrug-resistant Acinetobacter baumannii, as it can rapidly develop resistance to current antibiotics. This is the first study to investigate the synergistic effect of colistin/doripenem combination on the metabolome of A. baumannii. The metabolite levels were measured using LC-MS following treatment with colistin (2 mg/L) or doripenem (25 mg/L) alone, and their combination at 15 min, 1 hr and 4 hr (n = 4). Colistin caused early (15 min and 1 hr) disruption of the bacterial outer membrane and cell wall, as demonstrated by perturbation of glycerophospholipids and fatty acids. Concentrations of peptidoglycan biosynthesis metabolites decreased at 4 hr by doripenem alone, reflecting its mechanism of action. The combination induced significant changes to more key metabolic pathways relative to either monotherapy. Down-regulation of cell wall biosynthesis (via D-sedoheptulose 7-phosphate) and nucleotide metabolism (via D-ribose 5-phosphate) was associated with perturbations in the pentose phosphate pathway induced initially by colistin (15 min and 1 hr) and later by doripenem (4 hr). We discovered that the combination synergistically killed A. baumannii via time-dependent inhibition of different key metabolic pathways. Our study highlights the significant potential of systems pharmacology in elucidating the mechanism of synergy and optimizing antibiotic pharmacokinetics/pharmacodynamics

Abstracts of the International Halal Science Conference 2023

This book presents the extended abstracts of the selected contributions to the International Halal Science Conference, held on 22-23 August 2023 by the International Institute for Halal Research and Training (INHART), IIUM, Malaysia in collaboration with Halalan Thayyiban Research Centre, University Islam Sultan Sharif (UNISSA), Brunei Darussalam. With the increasing global interest in halal products and services, this conference is timely. Conference Title:  International Halal Science ConferenceConference Acronym: IHASC23Conference Theme: Halal Industry Sustainability Through ScienceConference Date: 22-23 August 2023Conference Venue: International Islamic University (IIUM), MalaysiaConference Organizer: International Institute for Halal Research and Training (INHART), International Islamic University (IIUM), Malaysi