Md. Imtaiyaz Hassan

Zinc α2-Glycoprotein: A Multidisciplinary Protein

Zinc α2-glycoprotein (ZAG) is a protein of interest because of its ability to play many important functions in the human body, including fertilization and lipid mobilization. After the discovery of this molecule, during the last 5 decades, various studies have been documented on its structure and functions, but still, it is considered as a protein with an unknown function. Its expression is regulated by glucocorticoids. Due to its high sequence homology with lipid-mobilizing factor and high expression in cancer cachexia, it is considered as a novel adipokine. On the other hand, structural organization and fold is similar to MHC class I antigen-presenting molecule; hence, ZAG may have a role in the expression of the immune response. The function of ZAG under physiologic and cancerous conditions remains mysterious but is considered as a tumor biomarker for various carcinomas. There are several unrelated functions that are attributed to ZAG, such as RNase activity, regulation of melanin production, hindering tumor proliferation, and transport of nephritic by-products. This article deals with the discussion of the major aspects of ZAG from its gene structure to function and metabolism. (Mol Cancer Res 2008;6(6):892–906)

Structure, function and applications of carbonic anhydrase isozymes.

The carbonic anhydrases enzymes (CAs, EC 4.2.1.1) are zinc containing metalloproteins, which efficiently catalyse the reversible conversion of carbon dioxide to bicarbonate and release proton. These enzymes are essentially important for biological system and play several important physiological and patho-physiological functions. There are 16 different alpha-carbonic anhydrase isoforms studied, differing widely in their cellular localization and biophysical properties. The catalytic domains of all CAs possess a conserved tertiary structure fold, with predominately β-strands. We performed an extensive analysis of all 16 mammalian CAs for its structure and function in order to establish a structure-function relationship. CAs have been a potential therapeutic target for many diseases. Sulfonamides are considered as a strong and specific inhibitor of CA, and are being used as diuretics, anti-glaucoma, anti-epileptic, anti-ulcer agents. Currently CA inhibitors are widely used as a drug for the treatment of neurological disorders, anti-glaucoma drugs, anti-cancer, or anti-obesity agents. Here we tried to emphasize how CAs can be used for drug discovery, design and screening. Furthermore, we discussed the role of CA in carbon capture, carbon sensor and metabolon. We hope this review provide many useful information on structure, function, mechanism, and applications of CAs in various discipline.

Luminol-Based Chemiluminescent Signals: Clinical and Non-clinical Application and Future Uses

Chemiluminescence (CL) is an important method for quantification and analysis of various macromolecules. A wide range of CL agents such as luminol, hydrogen peroxide, fluorescein, dioxetanes and derivatives of oxalate, and acridinium dyes are used according to their biological specificity and utility. This review describes the application of luminol chemiluminescence (LCL) in forensic, biomedical, and clinical sciences. LCL is a very useful detection method due to its selectivity, simplicity, low cost, and high sensitivity. LCL has a dynamic range of applications, including quantification and detection of macro and micromolecules such as proteins, carbohydrates, DNA, and RNA. Luminol-based methods are used in environmental monitoring as biosensors, in the pharmaceutical industry for cellular localization and as biological tracers, and in reporter gene-based assays and several other immunoassays. Here, we also provide information about different compounds that may enhance or inhibit the LCL along with the effect of pH and concentration on LCL. This review covers most of the significant information related to the applications of luminol in different fields.

Role of N-terminal residues on folding and stability of C-phycoerythrin: simulation and urea-induced denaturation studies

The conformational state of biliproteins can be determined by optical properties of the covalently linked chromophores. Recently determined crystal structure of truncated form of α-subunit of cyanobacterial phycoerythrin (αC-PE) from Phormidium tenue provides a new insight into the structure–function relationship of αC-PE. To compare their stabilities, we have measured urea-induced denaturation transitions of the full length αC-PE (FL-αC-PE) and truncated αC-PE (Tr-αC-PE) followed by observing changes in absorbance at 565 nm, fluorescence at 350 and 573 nm, and circular dichroism at 222 nm as a function of [urea], the molar concentration of urea. The transition curve of each protein was analyzed for ΔGD0, the value of Gibbs free energy change on denaturation (ΔGD) in the absence of urea; m, the slope (=∂∆GD/∂[urea]), and Cm, the midpoint of the denaturation curve, i.e. [urea] at which ΔGD = 0. A difference of about 10% in ΔGD0 observed between FL-αC-PE and Tr-αC-PE, suggests that the two proteins are almost equally stable, and the natural deletion of 31 residues from the N-terminal side of the full length protein does not alter its stability. Furthermore, normalization of probes shows that the urea-induced denaturation of both the proteins is a two-state process. Folding of both structural variants (Tr-αC-PE and FL-αC-PE) of P. tenue were also studied using molecular dynamics simulations at 300 K. The results show clearly that the stability of the proteins is evenly distributed over the whole structure indicating no significant role of N-terminal residues in the stability of both proteins.

Large scale analysis of the mutational landscape in β-glucuronidase: A major player of mucopolysaccharidosis type VII

The lysosomal storage disorders are a group of 50 unique inherited diseases characterized by unseemly lipid storage in lysosomes. These malfunctions arise due to genetic mutations that result in deficiency or reduced activities of the lysosomal enzymes, which are responsible for catabolism of biological macromolecules. Sly syndrome or mucopolysaccharidosis type VII is a lysosomal storage disorder associated with the deficiency of β-glucuronidase (EC 3.2.1.31) that catalyzes the hydrolysis of β-D-glucuronic acid residues from the non-reducing terminal of glycosaminoglycan. The effects of the disease causing mutations on the framework of the sequences and structure of β-glucuronidase (GUSBp) were analyzed utilizing a variety of bioinformatic tools. These analyses showed that 211 mutations may result in alteration of the biological activity of GUSBp, including previously experimentally validated mutations. Finally, we refined 90 disease causing mutations, which presumably cause a significant impact on the structure, function, and stability of GUSBp. Stability analyses showed that mutations p.Phe208Pro, p.Phe539Gly, p.Leu622Gly, p.Ile499Gly and p.Ile586Gly caused the highest impact on GUSBp stability and function because of destabilization of the protein structure. Furthermore, structures of wild type and mutant GUSBp were subjected to molecular dynamics simulation to examine the relative structural behaviors in the explicit conditions of water. In a broader view, the use of in silico approaches provided a useful understanding of the effect of single point mutations on the structure-function relationship of GUSBp.

Curcumin specifically binds to the human calcium–calmodulin-dependent protein kinase IV: fluorescence and molecular dynamics simulation studies

Calcium–calmodulin-dependent protein kinase IV (CAMK4) plays significant role in the regulation of calcium-dependent gene expression, and thus, it is involved in varieties of cellular functions such as cell signaling and neuronal survival. On the other hand, curcumin, a naturally occurring yellow bioactive component of turmeric possesses wide spectrum of biological actions, and it is widely used to treat atherosclerosis, diabetes, cancer, and inflammation. It also acts as an antioxidant. Here, we studied the interaction of curcumin with human CAMK4 at pH 7.4 using molecular docking, molecular dynamics (MD) simulations, fluorescence binding, and surface plasmon resonance (SPR) methods. We performed MD simulations for both neutral and anionic forms of CAMK4-curcumin complexes for a reasonably long time (150 ns) to see the overall stability of the protein–ligand complex. Molecular docking studies revealed that the curcumin binds in the large hydrophobic cavity of kinase domain of CAMK4 through several hydrophobic and hydrogen-bonded interactions. Additionally, MD simulations studies contributed in understanding the stability of protein–ligand complex system in aqueous solution and conformational changes in the CAMK4 upon binding of curcumin. A significant increase in the fluorescence intensity at 495 nm was observed (λexc = 425 nm), suggesting a strong interaction of curcumin to the CAMK4. A high binding affinity (KD = 3.7 × 10−8 ± .03 M) of curcumin for the CAMK4 was measured by SPR further indicating curcumin as a potential ligand for the CAMK4. This study will provide insights into designing a new inspired curcumin derivatives as therapeutic agents against many life-threatening diseases.

Current updates on computer aided protein modeling and designing

Determination of the three dimensional (3D) structure of a protein can provide important details about its biological functions and mechanism of action. However, despite their significance, the precise three-dimensional structures of most of the proteins are not fully determined till date. The main focus of the current review article is to gain a better understanding of the structural features of the proteins using computational techniques, and their relationship with function. Protein modeling and design is the method aimed to fold a primary amino acids sequence into protein structure with the ultimate goal of designing novel function and behavior. Moreover, proteins can also be designed from scratch or by similarity with the known protein structure. In the current article we have tried to cover various computer aided protein modeling and designing via homology and ab initio modeling, folding study using Molecular Dynamics (MD) methods and in silico mutation analysis.

Microtubule affinity-regulating kinase 4: structure, function, and regulation

MAP/Microtubule affinity-regulating kinase 4 (MARK4) belongs to the family of serine/threonine kinases that phosphorylate the microtubule-associated proteins (MAP) causing their detachment from the microtubules thereby increasing microtubule dynamics and facilitating cell division, cell cycle control, cell polarity determination, cell shape alterations, etc. The MARK4 gene encodes two alternatively spliced isoforms, L and S that differ in their C-terminal region. These isoforms are differentially regulated in human tissues including central nervous system. MARK4L is a 752-residue-long polypeptide that is divided into three distinct domains: (1) protein kinase domain (59–314), (2) ubiquitin-associated domain (322–369), and (3) kinase-associated domain (703–752) plus 54 residues (649–703) involved in the proper folding and function of the enzyme. In addition, residues 65–73 are considered to be the ATP-binding domain and Lys88 is considered as ATP-binding site. Asp181 has been proposed to be the active site of MARK4 that is activated by phosphorylation of Thr214 side chain. The isoform MARK4S is highly expressed in the normal brain and is presumably involved in neuronal differentiation. On the other hand, the isoform MARK4L is upregulated in hepatocarcinoma cells and gliomas suggesting its involvement in cell cycle. Several biological functions are also associated with MARK4 including microtubule bundle formation, nervous system development, and positive regulation of programmed cell death. Therefore, MARK4 is considered as the most suitable target for structure-based rational drug design. Our sequence, structure- and function-based analysis should be helpful for better understanding of mechanisms of regulation of microtubule dynamics and MARK4 associated diseases.

Protein aggregation and neurodegenerative diseases: From theory to therapy.

The study of protein misfolding and aggregation saw resurgence in the last decade. Aggregation is the main cause of several human neurodegenerative diseases which makes this field as the leading edge in the science today. Protein aggregation is a highly complex process resulting in formation of a variety of aggregates with different structures and morphologies. Many of them are highly cytotoxic. In-depth knowledge about structure, mechanism of formation, and physiological effects of aggregates will shed new light on the aggregation-mediated cell toxicity, and helps in deciphering new target for drug design and development. This review summarizes the existing information on the molecular mechanism of protein misfolding and aggregation involved in neurodegeneration stressing on the possible therapeutic intervention in neurodegenerative diseases. As our knowledge about the relation between the protein misfolding and disease pathogenesis will be enhanced, more specific and promising treatment opportunities will come into existence.

Therapeutic progress in amyotrophic lateral sclerosis-beginning to learning.

Amyotrophic lateral sclerosis (ALS) is a fatal neurodegenerative disease associated with motor neuron degeneration, muscle weakness, paralysis and finally death. The proposed mechanisms of ALS include glutamate excitotoxicity, oxidative stress, inflammation, mitochondrial dysfunction, apoptosis and proteasomal dysfunction. Although numerous pathological mechanisms have been explained, ALS remains incurable disease because of failure of clinical trials and lack of any effective therapy. The rapid advancement in genetic discoveries in ALS emphasizes the point that ALS is a multi-subtype syndrome rather than a single disease. This can be argued as one of the single reason why many previous therapeutic drug trials have failed. Efforts to develop novel ALS treatments which target specific pathomechanisms are currently being pursued. Herein, we review the recent discovery and preclinical characterization of neuroprotective compounds and compare their effects on disease onset, duration and survival. Furthermore, the structure-activity relationships of these agents are analyzed with the overall goal of developing a screening strategy for future clinical applications.