American Journal of Biochemistry and Molecular Biology - Current Issue

Homology Modeling of a Fruit Ripening Specific Plant MADS–box Factor

20 May, 2013

A MADS-box (Minichromosome maintenance-1, Agamous, Deficiens and Serum response factor) transcription factor namely SlMADS RIN (Solanum lycopersicum MADS Ripening inhibitor) seems to act as global regulator in climacteric fruit ripening process of tomato. Structure modeling of any plant MIKC (MADS-box, I-box, K-box and C-box) -type MADS-box factor were unknown till date, and the present study is an approach towards this direction. The template search of SlMADS RIN was performed by PSI BLAST (Position-Specific Iterative Basic Local Alignment Search Tool). Initial model was built with the help of MODELLER 9v4 package. The predicted 3D structure for SlMADS RIN protein was further validated by Ramachandran plot analysis using the PROCHECK tool. The submitted sequence of SlMADS RIN protein to PSI BLAST tool identified only one region (1-73 amino acids). DOPE (Discrete Optimized Protein Energy) score analysis revealed that the modeled structure showed overall lower DOPE score value (-3968.569336). Ramachandran plot analysis revealed that 94.1% residues were in favored region and 05.9% residues were in allowed region. Result revealed that the SlMADS RIN protein structure under study deviate largely by sequence with the known MADS-box, except the N-terminal 74 amino acid. Further, the side chain and loops of SlMADS RIN showed <1Å (0.233 Å) root mean square deviation. Thus, it can be concluded that this is the first report on prediction of three dimensional models for SlMADS RIN and this modeled structure can be used to predict the molecular function of the protein.

Oxidative Stress Biomarkers and Biochemical Profile in Broilers Chicken Fed Zinc Bacitracin and Ascorbic Acid under Hot Climate

20 May, 2013

Dietary manipulation of heat induced oxidative stress is one of the most widely used method to alleviate the negative effect of heat stress. The combination of two or more potential antioxidants are known to protect against heat stress. Therefore, 200 day old broiler chicks were raised under hot climate to determine the effect of ascorbic acid and zinc bacitracin and their combination as antioxidants. Birds were divided into four treatments; the first treatment (T₁), the control was provided with basal diet. Second (T₂) and third (T₃) treatments were provided with 1 g ascorbic acid and 100 mg zinc bacitracin per kg basal diet, respectively. The fourth treatment (T₄) was provided with a combination of T₂ and T₃ for 42 days. Performance parameters were reported every fortnight. Blood samples were collected every fortnight and the harvested sera were used to determine oxidative stress biomarkers. Ascorbic acid and zinc bacitracin were safe for heat stressed birds since the liver and kidney function indicators were not changed. Ascorbic acid and zinc bacitracin lowered lipid peroxidation level (Malondialdehyde; MDA) and increased the activity of catalase (CAT), glutathione-S-transferase (GST) and Super Oxide Dismutase (SOD) whereas, glutathione peroxidase (GPx) activity remained unchanged in all treatments. Serum ascorbic acid concentration was significantly (p<0.05) increased accompanied with hypocholesterolemia in heat stressed broilers receiving ascorbic acid compared to the control. Performance parameters were not influenced by feed additives. In conclusion, ascorbic acid was more potent than zinc bacitracin and their combination in increasing the level of enzymatic and non enzymatic antioxidants.

Modulatory Effect of Aqueous Stem Bark Extract of Psidium guajava Linn. against CCl4 Induced Liver Damage in Rats

20 May, 2013

The present study was aimed to evaluate the stem bark aqueous extract of Psidium guajava for modulatory effect against CCl₄ induced liver damage in rats. A total of thirty six male rats, were randomly divided into six groups of six rats each. The extract was administered orally for 15 days at 125, 250 and 500 mg kg^-1 b.wt. The results obtained showed that treatment with the extract significantly (p<0.05) restored liver weight. There was significant (p<0.05) increase in the level of Packed Cell Volume (PCV), haemoglobin (Hb) and Red Blood Cell (RBC) counts and significant (p<0.05) decrease in White Blood Cell (WBC) counts compared to toxin control group. Also administration of the extract caused significant (p<0.05) decrease in the activities of Alanine Transaminase (ALT), Aspartate Transaminase (AST), Alkaline Phosphatase (ALP) and the level of total bilirubin and significant (p<0.05) increase in total protein level compared to toxin control group. Similarly the extract caused a significant (p<0.05) increase in the activities of Catalase (CAT) and Superoxide Dismutase (SOD) and significant (p<0.05) decrease in reduced Glutathione (GSH) and Thiobarbituric Reactive Substances (TBARS) level compared to group 2 (toxin control group). The histopathological study indicated that treatment with the extract restored and regenerated hepatic cells compared to toxin control group. This study found that administration of aqueous stem bark extracts ameliorated hepatotoxicity induced by CCl₄ in rats.

Hypolidemic and Angiotensin-converting Enzyme Inhibitory Effect of Peristrophe bicalyculata (Retz.) Nees

20 May, 2013

The objective of this study was to investigate the effect of Peristrophe bicalyculata on lipid levels and activity of angiotensin-converting enzyme in kidneys, lungs and heart of fat-fed obese Wistar rats. The angiotensin-converting enzyme inhibition pattern of Peristrophe bicalyculata was also studied. Apparently healthy male Wistar rats were grouped into: control group, fat-fed control, standard control (atorvastatin at 70 mg kg^-1) and low dose (100 mg kg^-1) and high dose (250 mg kg^-1) extract groups, after confirmation of the lethal dose (LD₅₀). All rats, except those in the control group were fat-fed until obesity was confirmed (BMI>0.3) and treatment given for 4 weeks. From our results, serum and liver levels of Total Cholesterol (TC), Triacyglycerol (TG) and Low Density Lipoprotein increased significantly (p<0.05) in obese rats compared to the control rats; while High Density Lipoprotein (HDL) significantly decreased. Administration of atorvastatin and the extract significantly (p<0.05) reduced TC, TAG and LDL levels and increased HDL in serum and liver. The activity of Angiotensin Converting Enzyme (ACE) in kidneys, lungs and heart of obese rats was significantly (p<0.05) higher than that of rats in the control group. The extract reduced ACE activity in the lungs, kidneys and heart by 97, 98 and 96%, respectively, which was 20, 30 and 15 folds that of the standard drug. Studies on the inhibition pattern of the extract showed that Peristrophe bicalyculata inhibits ACE by competing with the substrate for the active site, exhibiting a competitive type of inhibition. The K_m of ACE increased from 0.25 to 2.5 mM at 1% concentration and 5 mM at 2% with an unchanged V_max and k_ivalue of 1.1 mg mL^-1. In conclusion, this study has confirmed that the aqueous extract of Peristrophe bicalyculata may be useful in the reduction of cardiovascular disease risk.

Ultrasonication of Chicken Natural Actomyosin: Effect on ATPase Activity, Turbidity and SDS-PAGE Profiles at Different Protein Concentrations

20 May, 2013

With the increasing application of ultrasonics in meat tenderization and processing, physicochemical events initiated by sonic radiation at myofibrillar level and propagated in complex tissue such as meat require a clear understanding. The enormous amount of basic information collected by studying myofibrils, actomyosin or their individual constituents has already clarified intricacies of muscular contraction and, part of such basic information has find application in meat sciences. In this investigation, chicken Natural actomyosin (NAM) has been taken as a simple model to work out some effects of ultrasonication in a concentration range of 0.5 to 1.8 mg protein mL^-1. At each concentration, NAM solution in 0.6 M NaCl (2.0 mL) was individually exposed to 20 kHz sonic waves for a total of 10 min. Cooling was maintained by keeping NAM containers in crushed ice and a lag of 5 sec after each 10 sec long sonic burst. Aliquots from each sonicated NAM were subjected to biochemical analyses. Most striking differences were observed in Ca²⁺-ATPase activity, which displayed a steady decline that corresponded with the decreasing protein concentration. Ultrasonication of NAM for 10 min caused a loss of ~47% of Ca²⁺-ATPase activity at the highest dilution (0.5 mg mL^-1). In the same order of protein concentration, turbidity of ultrasonicated NAM also decreased which denotes increasing transparency. Thus, ATPase and turbidity data demonstrate that due to sonic radiation, interactions among constituents of chicken actomyosin complex alter and these structural changes are devoid of any fragmentation. Under present experimental conditions, SDS-PAGE profiles did not reveal any novel band which could be attributed to ultrasonic fragmentation or proteolytic contamination. The findings also suggest that unlike myofibrils, actomyosin is a model where interactions and substructural changes of constituent polypeptides can be investigated without interference of endogenous muscle proteases.

Induction of Salicylic Acid in Cucumber Plants Against Cucumber Mosaic Cucumovirus Using Biotic Inducers

20 May, 2013

Systemic Acquired Resistance (SAR) could be induced in cucumber plants using different individual of seven microbial isolates against Cucumber mosaic Cucumovirus (CMV). These isolates were Bacillus subtilis, Bacillus polymyxa, Bacillus circulans, Pseudomonas putida, Pseudomonas fluorescens 2 and Pseudomonas fluorescens 8 as bacterial isolates and Trichoderma harzianum as fungal isolate. The occurrence of SAR was found by seed treatment with microbial liquid culture based on virus infectivity and the level of free and endogenous salicylic acid (SA), 14 days from CMV inoculation. Seven biotic inducers reduced CMV infectivity at range 16.6 to 39% and T. harzianum gave the highest percentage of reduction 39%. In addition, the level of total SA has been increased in treated plants, B. subtilis gave the highest level of SA 239.13 μg g^-1 fresh weight (fwt) while, B. circulans gave the lowest level 70.1 μg g^-1 fwt.

Partial Characterization of Proteolytic and Milk Clotting Enzymes in Sodom Apple Calotropis procera (Ait.) R.Br. (Asclepiadaceae) Plant

20 May, 2013

Calotropis procera (Sodom apple) or Bomubomu in Yoruba Language has been used locally in the Northern part of Nigerian and the Fulani set-ups in African settings as curdling agent in the production of ‘warankasi’ (a local soft cheese) when using cow milk; this plant is also known for its medicinal importance like antidote for snake bite, sinus fistula, rheumatism, mumps, burn injuries, body pain and treatment of jaundice in India and some parts of Africa. However, this present study was undertaken to characterize the milk clotting and proteolytic enzymes in the plant with the aim of establishing the optimal conditions of the enzymes to be used for cheese and allied milk products manufacturing and protein hydrolytic processes of medicinal importance. The results obtained from this study showed that the enzyme activities were evidenced in all the plant parts extract preparations except the root that had only proteolytic activity without noticeable milk-clotting activity for the period of one hour of investigation. The optimal pH for proteolytic and milk clotting enzyme activities were observed to be 4.0 for proteolytic and 5.0 for milk clotting enzyme activities in the crude extracts using casein and powdered milk as substrates, respectively while the temperature optima for the two enzyme activities were observed to be 70°C for milk clotting and 60°C for proteolytic. Calcium ion and cysteine were found to activate the two enzymes, with maximum activation at 800 mM Ca²⁺for proteolytic activity and 10 mM for milk clotting activity while cysteine showed maximum activation at concentration of 6.0 mM for the two enzymes as Pb²⁺ ions caused inhibition of both enzymes at the very least concentration. The results of analyzes from this work showed how the proteolytic and milk clotting enzyme activities of the plant latex could be enhanced for protein hydrolytic processes of medicinal importance and industrial production of cheese and allied milk products.

Detection of Canine Parvo Virus by Polymerase Chain Reaction Assay and its Prevalence in Dogs in and Around Mathura, Uttar Pradesh, India

20 May, 2013

The early detection of the Canine Parvo Virus (CPV) is of paramount importance. The present study was aimed to know the molecular epidemiology of Canine parvo virus. Canine faecal samples from 100 dogs showing the clinical signs of gastroenteritis in and around Mathura, Uttar Pradesh, India were collected and DNA was extracted by phenol-chloroform method. CPV vaccine strain was used as a positive control. Polymerase Chain Reaction (PCR) was carried out to amplify VPI/VP2 gene using a set of 20-mer primers [pCPV-RT (Forward): 5’-CAT TGG GCT TAC CAC CAT TT-3’; (Reverse): 5’-CCA ACC TCA GCT GGT CTC AT-3’)] from position 3136-3155 to 3276-3295 of VP1/VP2 gene. A PCR product of approximately 160 bp was generated with positive faecal samples and CPV vaccine strain. After screening, 63 dogs were found positive for CPV but no sex variation was noticed amongst the CPV positive cases. Dogs, of the age group of ≤6 months were more susceptible in comparison to of >6 months and highest occurrence was noted in unvaccinated dogs and dogs in co-habitation with other dogs. Breed wise distribution of CPV in dogs revealed that the prevalence of CPV was the highest in Doberman (77.78%), followed by Spitz (78.57%), German shepherd (70.00%), Labrador (68.75%), Pomeranian (45.45%). It is concluded that CPV is prevalent in the Mathura and nearby area and it is more common in pups of age less than 6 months old and more prevalent in German shepherd, Labrador and Pomeranian breeds of dog.