BIOCHEMISTRY, BIOMEDICINE & PHARMACEUTICS

Introduction to Pentose Phosphate Pathway The pentose phosphate pathway ( Hexose monophosphate pathway ) is the metabolic pathway that occurs in the cytosol and generates NADPH (nicotinamide adenine dinucleotide phosphate) which is utilized in various biosynthetic pathways. This pathway can be broadly classified into two phases i.e oxidative phase and non-oxidative phase.  In the oxidative stage of the pentose phosphate pathway, the glucose-6-phosphate is converted to five carbon ribulose-5 phosphate with generation of NADPH.  During the non-oxidative stage, the interconversion of pentose sugars and hexose sugars is catalyzed by isomerases and aldolases. The fate of these reversible steps depends on the anabolic and energy status of the cells.  The two primary functions of the pentose phosphate pathway are ; a) Provide NADPH (a coenzyme) for biosynthetic pathways b) Provide pentose sugars for nucleotides (DNA and RNA) synthesis Two phases of Pentose phosphat...

 To label mitochondria, cells are simply incubated with commercially available Mito Tracker® fluorescent dye such as tetramethylrhodamine methyl ester (TMRM), and tetramethylrhodamine ethyl ester (TMRE), Rhodamine 123 which passively diffuse across the plasma membrane and accumulate in active mitochondria. Once their mitochondria are labeled, the cells can be treated with an aldehyde-based fixative for samples that need fixation to allow further processing of the sample. Some of the Mito Tracker® probes are also retained after permeabilization with some detergents during subsequent processing steps (e.g., immunocytochemistry or in situ hybridization). MitoTracker Red CMXRos MitoTracker Red CMXRos is a red-fluorescent dye that stains mitochondria in live cells and its accumulation is dependent upon membrane potential. The dye is well-retained after aldehyde fixation. allowing for further sample processing and immunostaining. Excitation: 579 nm, Emission: 599 nm,  Invitroge...

       Acid capture elution (ACE) is a plate-based method for reducing the drug interference in the ADA (Adenosine deaminase) assay. It is used for the purification and isolation of specific nucleic acids, such as DNA or RNA, from a complex mixture. This method utilizes weak organic acids (Glycine, Acetic acid) to dissociate the drug-ADA complex.  The sample is neutralized and allowed the bind the plate coated with the drug. After the ADA is captured, the plate is washed to remove any unbound drug present in the sample.  The second acid treatment is used to elute the ADA bound to the plate.  The ADA sample is transferred, neutralized with Tris-HCl base, and processed in the ADA detection assay.  Procedure: 1) Binding (Coating of the plate with a drug): Maxisorp ELISA Nunc plate is widely used for coating the plate that would be used for the capture of ADA. For the optimization of drug concentration, test different concentrations of drug (0....

Heme Biosynthesis and Regulation Biological Importance of Heme  - Oxygen transport (Hemoglobin) & Storage (Myoglobin) - Electron carrier in Electron Transport Chain  (Cytochromes) - Prosthetic group of various enzyme - Deficiency of the enzyme causes porphyria Biosynthetic Pathway for the Synthesis of Heme  Figure 1: Pathway for Heme Synthesis (Source: NEJM).   Steps of the pathway: 1. Formation of Delta Aminolevulinic Acid (ALA): - catalyzed by enzyme ALA synthase, rate limit step for the biosynthesis of Heme - ALA synthase (ALAS) is a mitochondrial enzyme consisting of two isozymes - ALAS1 is a housekeeping gene present in the cells consisting of mitochondria - ALAS2 is the erythroid form of the ALA synthase - Succinyl CoA and Glycine are the precursors that undergo condensation to form ALA - ALAS require pyridoxal-5-phosphate as a cofactor - The deficiency of ALA synthase causes X-Linked Sideroblastic Anemia  2. Conversion of ALA to Porphobilinogen (...

Q) Which of the following is a FALSE statement about the fate of the glycolytic pathway in erythrocytes? a) Glycolysis is the sole source of ATP (adenosine triphosphate) in erythrocytes. b) Deficiency of pyruvate kinase leads to hemolytic anemia. c) Pyruvate generated during glycolysis is converted to lactate. d) Pyruvate generated during glycolysis is converted to acetyl CoA and enters the TCA cycle (tricarboxylic acid cycle) in mitochondria. (Correct Answer: Please find it at the end of this text ) The objective of the MCQ above is to discuss - Shape and morphology of   Red blood cell - The fate of glucose in red blood cells - Metabolic control of glucose metabolism Red blood cells are bone marrow-derived non-dividing cells that contain hemoglobin ( 95% of intracellular protein ) and help to transport oxygen from the lungs to peripheral tissue.  The RBCs help in the disposal of carbon dioxide and a proton from the peripheral tissue, they are biconcave in the s...

History of Gene Therapy, Vector DNA integration, and Safety concern: After more than two decades of setbacks and scrutiny, gene therapy has reemerged in the mainstream of drug discovery. This has renewed hope and enthusiasm among patients, healthcare providers, and drug developers.  During early clinical trials, there were incidences of life-threatening safety risks associated with gene therapy and death, these events brought the human trials near to a halt.  Genotoxicity, cancer induction, and progression were common safety events observed in numerous clinical trials.   It was believed that these gene therapy products may have induced  activation of protooncogene, clonal expansion of these tumorigenic cells, and progression of leukemia. The viral vector DNA insertion into the host DNA may be associated with the mutation resulting in various tumor-promoting phenomena. Viral Vectors, Gene Integration, and Integration Assay: Lentivirus , Gamma retroviruses , a...

                Multiple Choice Question on Pseudomonas spp & Acinetobacter spp Gram positive or gram negative bacteria? Cocci or bacilli? 1) Which of the following is not the general characteristic of Pseudomonas spp ? a) Rod-shaped, curved b) Motile c) Found in soil and water and plants d) All species cause diseases in humans only 2) Pseudomonas aeruginosa is capable of producing two water-soluble pigments that contribute to the virulence of the pseudomonas infections, one is blue color pigment called pyocyanin and …....................... color pigment called pyoverdin. a) Green b) Blue-green c) Red d) Pink-red 3) Patients with cystic fibrosis infection suffer from a chronic lung infection caused by Pseudomonas aeruginosa .  The bacterial growth results in the formation of ................................ in the host cells. a) Pigment b) Biofilm c) White blood cells d) Endotoxin 4) Which of the following temper...

Lecture Notes and Multiple Choice Question  on Inheritance Pattern of Genetic Disorders Multiple Choice Questions on Inheritance Pattern of Genetic Disorders 1) Which of the following statement is true regarding the Mendelian Theory of Inheritance?  a) The genotype ratio for the second generation is 1:2:1 b) The phenotype ratio for the second generation is 3:1 c) The inheritance pattern assume one gene-one protein phenomenon d) All of the above 2) Which of the following statements are true regarding the dominant traits? a) One dominant allele ( heterozygotes ) is sufficient to show the dominant trait b) Both dominant alleles  ( homozygous ) are required to show the dominant trail c) Heterozygotes ( individuals with one allele ) are generally carriers d) None of the Above 3) Which of the following statements are true regarding the recessive traits? a) One recessive allele (heterozygotes) is sufficient to show the phenotypic trait b) Both recessive alleles  (homoz...

                    MCQ on Circulatory Pathology : Clinical Pathology 1) Edema is the presence of excess fluid in the intracellular space.  The increased hydrostatic pressure causes edema in which of the following condition? a) Congestive failure b) Portal Hypertension c) Venous Thrombosis d) All of the above e) None of the above 2) Which of the following condition causes edema with hypoalbuminemia & decreased colloid osmotic pressure? a) Nephrotic syndrome b) Primary hyperaldosteronism  c) Renal Failure d) Graves disease 3) Which of the following disease is associated edema caused by increase endothelial permeability? a) Hypersensitivity reaction type I b) Bleomycine treatment c) Renal failure d) Graves disease 4) Exudate is the type of edema with high protein content and cells.  Which of the following are true for exudate ? a)  Results from increased vascular permeability caused by inflammation b) Specific gravi...

Overview of the Topics Anticoagulant Drugs Thrombolytic Drugs (Click Here) Antiplatelet Drugs (Click Here) Multiple Choice Practice Questions (Click Here) Anticoagulant drugs:  - Drugs that inhibit the development and enlargement of clots by action on the coagulation phase.  - Do not lyse clots or affect the fibrinolytic pathways  Clinical Indication for Anticoagulant therapy - Deep Vein Thrombosis - Arterial Embolism - Arterial Fibrillation  - Unstable angina and myocardial infarction Heparin (Intravenous or subcutaneous administration) - Heparin is a sulfated acidic mucopolysaccharide that is widely distributed in the body - Two types of heparin are used clinically - Standard Unfractionated Heparin - Low Molecular Weight Heparin Standard Unfractionated Heparin - Heparin (heparin sodium) is a mixture of highly electronegative acidic mucopolysaccharides that contain numerous N- and O-sulfate linkages.  - The unfractionated preparation is extracted f...

                 MCQ on Immunodeficiency diseases/Immunodeficiency disorders Immunodeficiency diseases or disorders: It refers to the disorders characterized by a weakened or impaired immune system. These disorders leads an individual more susceptible to infections and other diseases because their immune system is unable to adequately defend against microbial infections. 1. Which of the following is a rare and severe form of immunodeficiency where individuals lack T cells and B cells? a) DiGeorge Syndrome b) Wiskott-Aldrich Syndrome (WAS) c) Severe Combined Immunodeficiency (SCID)     d) Common Variable Immunodeficiency (CVID) 2. Wiskott-Aldrich Syndrome (WAS, an X-linked disorder) is associated with which of the following condition? a) Hypertension    b) Asthma  c) Diabetes mellitus  d) Eczema       3. Which of the following is not a primary immunodeficiency disorder?...

Note: MCQs with answers at the end  Hemoglobin Structure : - Hemoglobin is a heme-containing protein of the red blood cells - Globular proteins with a diameter of 6.4 nm with a molecular weight of 64.5 kDa - Heterodimers consisting of two  α  and two    β  subunits - Each subunit consist of a heme pocket that binds to the oxygen - Heme is a tetrapyrrole ring structure consisting of  Fe+2 at the center - The  α  subunit consist of 141 amino acids and the  β  subunit consists of 146 amino acids - The hydrophobic interaction of the  α β  dimers tightly held the quaternary structure of hemoglobin Hemoglobin Function -   Hemoglobin is present in the Red Blood Cells (RBCs) and primarily carries oxygen from the lungs to the peripheral tissues - One hemoglobin binds to four oxygen molecules and shows cooperativity - Binding of one oxygen to a subunit induces the conformational change to convert T-state to R-state -The R-state...