Abu Bakar Salleh

Malaysia Genome Institute, Kuala Lumpor, Kuala Lumpur, Malaysia

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Publications (283)438.62 Total impact

  • M. Basri · R.N.Z.R. Abd Rahman · A.B. Salleh
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    ABSTRACT: Palm-based specialty oleochemicals are special industrial chemicals from palm oil which are highly priced with high profit margins. These oleochemicals exhibit interesting properties such as excellent emolliency, surface activity, emulsifying properties as well as beneficial biological properties. As such, these compounds find many applications in the cosmetic, pharmaceutical and food industries. Enzyme catalysed syntheses of these chemicals are preferable as compared to their usual petro-chemical counterpart as these processes are nature identical and 'green'. Lipase-catalysed syntheses of specialty oleochemicals were carried out. The oleochemicals include amino acid esters, fatty alkanolamides, fatty esteramines, various wax esters, medium chain triglycerides and biologically active esters. The optimisation of the reaction conditions was discussed. The effects of the parameters influencing the reactions including temperature, reaction time, substrate molar ratio, amount of enzyme and solvents used were presented. The characteristics of the reaction system and the products were determined.
    No preview · Article · Apr 2013 · Journal of oil palm research
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    ABSTRACT: The thermostable alkaline protease from Bacillus stearothermophilus F1 has high potential for industrial applications, and attempt to produce the enzyme in yeast for higher yield was undertaken. Secretory expression of F1 protease through yeast system could improve enzyme's capability, thus simplifying the purification steps. Mature and full genes of F1 protease were cloned into Pichia pastoris expression vectors (pGAPZ alpha B and pPICZ alpha B) and transformed into P. pastons strains (GS115 and SMD1168H) via electroporation method. Recombinant F1 protease under regulation constitutive GAP promoter revealed that the highest expression was achieved after 72 h cultivation. While inducible AOX promoter showed that 0.5% (v/v) methanol was the best to induce expression. It was proven that constitutive expression strategy was better than inducible system. The a-secretion signal from the plasmid demonstrated higher secretory expression level of F1 protease as compared to native Open Reading Frame (ORF) in G5115 strain (GE6GS). Production medium YPTD was found to be the best for F1 protease expression with the highest yield of 4.13 U/mL. The protein was expressed as His-tagged fusion protein with a size about 34 kDa.
    No preview · Article · Apr 2013 · Genes & Genetic Systems

  • No preview · Article · Mar 2013
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    ABSTRACT: Fatty acid esters are long-chain esters, produced from the reaction of fatty acids and alcohols. They possess potential applications in cosmetic and pharmaceutical formulations due to their excellent wetting behaviour at interfaces and a non-greasy feeling when applied on the skin surfaces. This preliminary work was carried out to construct pseudo-ternary phase diagrams for oleyl laurate, oleyl stearate and oleyl oleate with surfactants and piroxicam. Then, the preparation and optimization study via 'One-At-A-Time Approach' were carried out to determine the optimum amount of oil, surfactants and stabilizer using low-energy emulsification method. The results revealed that multi-phase region dominated the three pseudo-ternary phase diagrams. A composition was chosen from each multi-phase region for preparing the nanoemulsions systems containing piroxicam by incorporating a hydrocolloid stabilizer. The results showed that the optimum amount (w/w) of oil for oleyl laurate nanoemulsions was 30 and 20 g (w/w) for oleyl stearate nanoemulsions and oleyl oleate nanoemulsions. For each nanoemulsions system, the amount of mixed surfactants and stabilizer needed for the emulsification to take place was found to be 10 and 0.5 g (w/w), respectively. The emulsification process via high-energy emulsification method successfully produced nano-sized range particles. The nanoemulsions systems passed the centrifugation test and freeze-thaw cycle with no phase failures, and stable for 3 months at various storage temperatures (3°C, 25°C and 45°C). The results proved that the prepared nanoemulsions system cannot be formed spontaneously, and thus, energy input was required to produce nano-sized range particles.
    No preview · Article · Feb 2013 · AAPS PharmSciTech
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    ABSTRACT: Since the industrial uses of lipase is expanding, research and development on this enzyme is strategically focusing on modifying its chemical, physical and biological properties. Surface residues of enzymes are reactive chemically and thus a potential target for chemical modification. We have carried out chemical modification via reductive alkylation on lysine residues of T1 and Candida rugosa lipase (CRL) using propionaldehyde. Through molecular dynamics (MD) and circular dichroism (CD), intrinsic and extrinsic fluorescence studies, the altered properties of T1 lipase were examined. Reductive alkylation using propionaldehyde caused the unfolding of enzymes as observed in chemically modified Candida rugosa and T1 lipases. As the effect of reductive alkylation is not localized at the modified site, the formation of molten globule could be observed in modified T1 lipase.
    No preview · Article · Jan 2013
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    ABSTRACT: Attractive applications of thermostable lipases in a vast array of industries are the driving forces for molecular biologists particularly to conduct more research on these enzymes. The advents in molecular biology techniques have now facilitated the researchers to manipulate genetic components of microorganisms hence producing the enzyme of interest in massive quantity with considerable cost and time saving. This chapter covers the molecular cloning and expression of new lipase genes into prokaryotic and eukaryotic systems, purification and characterization of the lipases. The lipase producing strains harbouring thermostable lipase genes were isolated from various geographical areas such as palm oil mill effluent, hot spring and oil contaminated area in Malaysia. Gene encoding for thermostable lipases were cloned and expressed in corresponding Escherichia coli and Pichia pastoris under the regulation of various strong promoters. These orthologous lipases share conserved functional motifs and 55-99% identities with molecular weight and pI about 43 kDa and 5.77-6.65, respectively. High yield purification (51-79%) could easily be achieved using single or two step purification strategies. T1 lipase, L2 lipase and L42 lipase optimally catalyzed triglycerides range from C8-C16. The thermal stability of T1 lipase was the highest among these lipases, while ARM lipase and L42 lipase were solvent tolerant lipases.
    No preview · Article · Jan 2013
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    ABSTRACT: This chapter discusses an overall view of protein crystallization of enzymes mainly on its basic purpose towards structure elucidation of a protein of interest. The existence of a protein crystal coupled with computer modelling gives the closest prescription in predicting the exact structure of the protein. We have successfully crystallized T1 lipase and its mutants; F16L and D311E lipases, L2 lipase and L42 lipase at 20°C, within 24 h to 14 d of incubation. The crystallization of T1 lipase and its mutants was carried out using vapour diffusion method, while counter diffusion method was applied for L2 and L42 lipases. T1 lipase was found to be successfully crystallised at various temperatures up to as high as 60°C, and this is the first report of protein crystallization at high temperature. By using these high quality crystals, the 3-D crystal structures of T1 lipases and its mutants as well as L2 lipase were successfully elucidated by X-ray crystallography. Furthermore, based on the crystal structure of T1 lipase, novel cation-P{cyrillic} interaction which formed between electron rich-aromatic ring and organic and metallic cations was found to exist. The studies of T1 lipase mutant D311E showed an improvement of thermal stability via outer-loop ion pair interaction.
    No preview · Article · Jan 2013
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    ABSTRACT: Rapid expansion of techniques in the field of molecular biology has made it possible to produce proteins especially enzymes from thermophilic microorganisms in mesophilic type. This will aid in cheaper, easier and perhaps abundant production of thermostable enzyme. Thermostable alkaline proteases are of great interest in the detergent industry because these enzymes are stable and able to retain its activities over a broad range of pH and temperatures. The gene for alkaline serine protease from thermophilic Bacillus stearothermophilus strain F1 was identified. Nucleotide sequence analysis showed an open reading frame of 1206 bp, coding for 401 amino acids. The deduced peptide sequence showed that F1 protease is produced from a pre-(N)pro-protease precursor consisting of 25, 97 and 279 amino acid residues respectively. The mature domain exhibited high similarity to Ak1 protease (97 %). The gene was expressed and excreted from the Escherichia coli XL1-Blue cytoplasm into the culture medium utilising the bacteriocin-release protein (BRP) system. The transformant harbouring two expression plasmids; the pTrcHis bearing F1 protease gene as well as pJL3 carrying BRP gene. The optimum conditions for the transformant to express and release F1 protease into the culture medium are discussed. The mature F1 protease with the size of 27 kDa was successfully released into the culture medium by E. coli transformant.
    No preview · Article · Jan 2013
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    ABSTRACT: Solvent tolerant microbes are able to grow in the presence of high concentrations of organic solvents, known to have detrimental effect on most microorganisms. The ability of these microbes to function in solvent-rich media is though their novel adaptation mechanisms as well as secretion of solvent stable enzymes. Our research group isolated several benzene-toluene-ethylbenzene-xylene (BTEX) tolerant bacteria locally. We have demonstrated the properties of the organic solvent tolerant Pseudomonas aruginosa S5 lipase, Bacillus sphaericus strain 205y and Stapylococcus epidermidis AT2. Molecular cloning and expression of new lipase genes of strain 205y and AT2 were achieved in PCR amplification method and expressed in pBAD TOPO TA and pTrcHis2, respectively. Most recently, we reported on the expression of a new intracellular organic solvent tolerant S5 lipase gene using a shot-gun cloning approach, after several attempts with PCR cloning failed. Previous failures might have been due to the fact that the Pseudomonas S5 lipase gene needs a helper or activator gene in the E. coli systems.
    No preview · Article · Jan 2013
  • M.S.M. Ali · R.N.Z.R.A. Rahman · N. Alias · A.B. Salleh · M. Basri
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    ABSTRACT: Psychrophilic organisms produce enzymes adapted to function at low temperature. These enzymes are characterized by a catalytic efficiency at low and moderate temperatures but are rather thermolabile. Thermophilic enzymes usually feature in discussion of industrial uses because their heat-stability makes them ideal biocatalyst for many reactions. However, cold-active/heat-labile enzymes also have great potential in industry. Cold-active enzymes might offer novel opportunities for biotechnological exploitation based on their high catalytic activity at low temperatures, low thermostability and unusual specificities. These properties are of interest in diverse fields such as detergents, fabric and food industry, bioremediation and biocatalysts under low water conditions. Furthermore, fundamental issues concerning the molecular basis of cold activity and the interplay between flexibility and catalytic efficiency are of important in the study of structure-function relationships in protein. By means of X-ray crystallography, these properties are beginning to be understood, and the rules governing their adaptation to cold appear to be relatively diverse. To date, extensive studies on psychrophilic enzymes were conducted ranging from purification and characterization, molecular cloning and expression, homology modeling and structural studies and x-ray crystallography studies. Lipases and protease which represents the most important groups of extracellular hydrolytic enzymes were examined due to the attractive properties of these enzymes that constitute a tremendous potential for fundamental research and biotechnological applications.
    No preview · Article · Jan 2013
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    ABSTRACT: This chapter is written to provide information on the production of heterologous proteins by yeast expression system, mainly by methylotrophic yeast Pichia pastorisand our work that utilizes P. pastoris to express thermostable lipases.Our attempt in isolating our own local yeast to be further developed as an expression hostwas also described. We have cloned and expressed thermostable L2 lipase in P. pastoris using AOX- and GAPderived promoter system, as well as tested fifth grade molasses to substitute glucose as potential carbon source. Our study showed that the use of fifth grade molasses was proven successful; however, further treatment on the molasses wasrequired to increase the amount of sugar content and to remove impurities. Cultivation of recombinant L2 lipase in bioreactor system with optimized conditionsof 35% air saturation, initial pH 6.0, 30 h of inoculum age, and 2% [w/v] of fifth grade molasses concentration produced 120 U/mL of lipase activity. This study showed that the constitutive pGAP expression system of P. pastoris is inducible by various carbon sources; therefore, it offers flexibility for diverse expressions of heterologous proteins. The activity of AT2 lipase expressed in Yarrowia lypolitica was successfully increased by 2.8-fold compared to Escherichia coliand 45-fold compared to wild types. Our study on isolation of local yeast to be developed as local host also resulted in discovery of potential new yeast species.
    No preview · Article · Jan 2013
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    ABSTRACT: Protein folding in enzymes and their sequences are not fully optimized, and only execute the minimum requirements in terms of folding efficiency and stability that permit proper functioning in the cell. There is still plenty of room for improvement through engineering of their structure and folding, whereby knowledge of structural features plays an important role. So far, significant structural data have been presented and the availability an abundance of structural data is the proponent of structure manipulations of enzymes. Also, the rapid progression in bioinformatics has resulted in the sudden surge of interests in protein structural analysis with the advent of innumerable tools. This chapter represents an overview of the various approaches taken in the rational design of T1 lipase of Geobacillus zalihae, ARM lipase of Geobacillus sp. and F1 protease of Bacillus stearothermophilus, to improve their stability under various conditions. Substitution on both oxyanion of T1 lipase have resulted in mutant lipases with increased thermostability, altered substrate specificity, varying stability toward different chemicals and altered enantioselectivity. Modification on the inter-loop of the helix lid of T1 lipase afforded mutant D311E with enhanced improved thermostability. The use of concensus approach by comparison with other homologous thermostable lipases helped identify the critical residue in ARM lipase. The R157S mutant of ARM lipase showed better compactness and increased the protein folding as opposed to the native lipase. Meanwhile, structural comparison of the F1 protease with other thermostable serine proteases had identified that the W200 as the hot residue to be replaced with Arg to increase the ion pair network in the F1 protease. Substitution of Arg afforded the W200R mutant with improved thermostability over the wild-type recombinant F1 protease.
    No preview · Article · Jan 2013
  • R.N.Z.R.A. Rahman · A.B. Salleh · M. Basri
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    ABSTRACT: This book serves as a tool to expand the knowledge of researchers in the discipline of microbiology and biochemistry encompassing the fields of molecular biology, structural biology, and crystallography for the two very important industrial enzymes: proteases and lipases. In addition it covers the fermentation technology for the production of locally isolated microbial lipases and proteases. This book presents work done in our laboratory which extensively reviews two major types of lipases and proteases: thermostable and/or organic-solvent tolerant including a chapter on cold-active and organic solvent tolerant lipase and protease isolated from Antarctica. Detailed descriptions on the isolation methods, protein production, protein characterization, and molecular cloning are discussed. Further works on structural biology of both proteases and lipases which include docking, molecular modification as well as protein crystallization are also presented. For protein crystallographers, this book highlights a novel discovery of high temperature crystallization of lipase. In addition, this book also offers a chapter on the cultivation strategy of lipase and protease in yeast expression systems and its bio-reactor studies. In short, this book is an all-in-one quick source of reference for researchers who want to have a quick grasp of understanding on many aspects of enzyme production, starting from the isolation and expression of the enzyme until crystallization and structure elucidation of the protein.
    No preview · Book · Jan 2013
  • C.F. Wong · R.N.Z.R.A. Rahman · A.B. Salleh · M. Basri
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    ABSTRACT: Proteases possess characteristics which make them desirable for industrial employments. Being present enormously in nature, proteases are obtainable from a variety of sources. This chapter reviews the necessities of utilizing organic-solvent tolerant proteases in industrial applications as well as the isolation works we have conducted to acquire the gene of interest from prospective microorganisms which naturally secrete proteases. New molecular cloning technology and expression in multiple hosts are highlighted in this chapter. Here, we present three proteases discovered from Pseudomonas aeruginosa strain K and Bacillus pumilus 115b. The proteases secreted by these microorganisms were proven stable in most of the tested organic solvents. The characteristics and behavior of elastase strain K in organic solvents are further discussed from the view of alteration in secondary structures, by subjecting methanol as the model.
    No preview · Article · Jan 2013
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    ABSTRACT: Chemical penetration enhancers such as micelles can be formed from the solubilisation of the nonionic surfactants in the nanoemulsion system which may then provide an effective solution to the problems associated with drug delivery systems. The structural properties of micelles comprising 5, 20, and 30 molecules of the nonionic surfactant Tween80 were investigated by all-atom level molecular dynamics simulation in explicit water system. The micelles with higher aggregation numbers showed a prolate shape, with the surface being dominated by the polar hydrophilic moieties of surfactant molecules. The average radius of gyration was between 1.1-4.9 nm while the effective radius, determined from the radius of gyration was between 1.4-3.2 nm. The estimated structural properties of the micelles formed may give us a more detail insight in understanding the complexity of these classes of nonionic surfactants.
    Full-text · Article · Dec 2012
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    ABSTRACT: Oil-in-water (O/W) nanoemulsions play an important key role in transporting bioactive compounds into a range of cosmeceutical products to the skin. Small droplet sizes have an inherent stability against creaming, sedimentation, flocculation, and coalescence. O/W emulsions varying in manufacturing process were prepared. The preparation and characterization of O/W nanoemulsions with average diameters of as low as 62.99 nm from palm oil esters were carried out. This was achieved using rotor-stator homogenizer and ultrasonic cavitation. Ultrasonic cell was utilized for the emulsification of palm oil esters and water in the presence of mixed surfactants, Tween 80 and Span 80 emulsions with a mean droplet size of 62.99 nm and zeta potential value at -37.8 mV. Results were comparable with emulsions prepared with rotor-stator homogenizer operated at 6000 rpm for 5 min. The stability of the emulsions was evaluated through rheology measurement properties. This included non-Newtonian viscosity, elastic modulus G', and loss modulus G″. A highly stable emulsion was prepared using ultrasonic cavitation comprising a very small particle size with higher zeta potential value and G' > G″ demonstrating gel-like behavior.
    No preview · Article · Dec 2012 · Journal of cosmetic science
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    ABSTRACT: Background Rheumatoid arthritis (RA) is an autoimmune disease with unknown etiology. Anticitrullinated protein autoantibody has been documented as a highly specific autoantibody associated with RA. Protein arginine deiminase type 4 (PAD4) is the enzyme responsible for catalyzing the conversion of peptidylarginine into peptidylcitrulline. PAD4 is a new therapeutic target for RA treatment. In order to search for inhibitors of PAD4, structure-based virtual screening was performed using LIDAEUS (Ligand discovery at Edinburgh university). Potential inhibitors were screened experimentally by inhibition assays. Results Twenty two of the top-ranked water-soluble compounds were selected for inhibitory screening against PAD4. Three compounds showed significant inhibition of PAD4 and their IC50 values were investigated. The structures of the three compounds show no resemblance with previously discovered PAD4 inhibitors, nor with existing drugs for RA treatment. Conclusion Three compounds were discovered as potential inhibitors of PAD4 by virtual screening. The compounds are commercially available and can be used as scaffolds to design more potent inhibitors against PAD4.
    Full-text · Article · Dec 2012 · BMC Bioinformatics
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    ABSTRACT: The crystallization of proteins makes it possible to determine their structure by X-ray crystallography, and is therefore important for the analysis of protein structure-function relationships. L2 lipase was crystallized by using the J-tube counter diffusion method. A crystallization consisting of 20% PEG 6000, 50 mM MES pH 6.5 and 50 mM NaCl was found to be the best condition to produce crystals with good shape and size (0.5 × 0.1 × 0.2 mm). The protein concentration used for the crystallization was 3 mg/mL. L2 lipase crystal has two crystal forms, Shape 1 and Shape 2. Shape 2 L2 lipase crystal was diffracted at 1.5 Å and the crystal belongs to the orthorhombic space group P2(1)2(1)2(1), with unit-cell parameters a = 72.0, b = 81.8, c = 83.4 Å, α = β = γ = 90°. There is one molecule per asymmetric unit and the solvent content of the crystals is 56.9%, with a Matthew's coefficient of 2.85 Å Da(-1). The 3D structure of L2 lipase revealed topological organization of α/β-hydrolase fold consisting of 11 β-strands and 13 α-helices. Ser-113, His-358 and Asp-317 were assigned as catalytic triad residues. One Ca(2+) and one Zn(2+) were found in the L2 lipase molecule.
    Full-text · Article · Dec 2012 · International Journal of Molecular Sciences
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    ABSTRACT: Palm oil-based esters (POEs) are unsaturated and non-ionic esters with a great potential to act as chemical penetration enhancers and drug carriers for transdermal drug nano-delivery. A ratio of palmitate ester and nonionic Tween80 with and without diclofenac acid was chosen from an experimentally determined phase diagram. Molecular dynamics simulations were performed for selected compositions over a period of 15 ns. Both micelles showed a prolate-like shape, while adding the drug produced a more compact micellar structure. Our results proposed that the drug could behave as a co-surfactant in our simulated model.
    Full-text · Article · Dec 2012 · International Journal of Molecular Sciences
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    ABSTRACT: Here, we describe an improved enzyme-facilitated epoxidation of 1-nonene using a conventional water bath shaker at ambient temperature. Enzymes were used to produce peroxy acids instantly from hydrogen peroxide (H2O2) and various perhydrolysis substrates. The peroxy acid generated was then utilised directly for in-situ oxidation of 1-nonene to 1-nonene oxide. Various parameters affecting the reaction were studied such as the nature of the peroxy acids, organic solvents, enzyme sources and enzyme concentrations. The highest conversion rate was achieved using phenylacetic acid as an oxygen carrier. 1-Nonene was converted most efficiently with 95% of the maximum yield by Novozym 435, an immobilised Candida antarctica lipase B, using dichloromethane as the reaction media. A minimum amount (16 mg, 1.4% w/w) of Novozym 435 was needed to maintain catalytic activity (160.0 Ug−1). In addition, a simple and rapid Gas chromatography mass spectroscopy selective ion monitoring (GC-MS SIM) method was developed using a HP-5ms column for determining 1-nonene oxide. The method was found to be linear in the range of 29.9 to 298.8 mg/L with R2 = 0.9981.
    No preview · Article · Nov 2012 · Biocatalysis and Biotransformation

Publication Stats

3k Citations
438.62 Total Impact Points

Institutions

  • 2012-2015
    • Malaysia Genome Institute
      Kuala Lumpor, Kuala Lumpur, Malaysia
  • 1999-2015
    • Putra University, Malaysia
      • • Faculty of Biotechnology and Biomolecular Sciences
      • • Department of Biochemistry
      • • Faculty of Environmental Studies
      Putrajaya, Putrajaya, Malaysia
  • 2003
    • National University of Malaysia
      • Department of Electrical, Electronic and Systems Engineering
      Putrajaya, Putrajaya, Malaysia
  • 1995
    • UPM
      Helsinki, Uusimaa, Finland