Biomedical subjects

Sankaran Krishnaswamy

Publications and source records attributed to Sankaran Krishnaswamy.

6 recordsLinked to original sources

PubMed2004

A simulation model of Escherichia coli osmoregulatory switch using E-CELL system.

BACKGROUND: Bacterial signal transduction mechanism referred to as a "two component regulatory systems" contributes to the overall adaptability of the bacteria by regulating the gene expression. Osmoregulation is one of the well-studied two component regulatory systems comprising of the sensor, EnvZ and the cognate response regulator, OmpR, which together control the expression of OmpC and OmpF porins in response to the osmolyte concentration. RESULTS: A quantitative model of the osmoregulatory switch operative in Escherichia coli was constructed by integrating the enzyme rate equations using E-CELL system. Using the substance reactor logic of the E-CELL system, a total of 28 reactions were defined from the injection of osmolyte till the regulated expression of porins by employing the experimental kinetic constants as reported in literature. In the case of low osmolarity, steady state production of OmpF and repression of OmpC was significant. In this model we show that the steady state - production of OmpF is dramatically reduced in the high osmolarity medium. The rate of OmpC production increased after sucrose addition, which is comparable with literature results. The relative porin production seems to be unaltered with changes in cell volume changes, ATP, EnvZ and OmpR at low and high osmolarity conditions. But the reach of saturation was rapid at high and low osmolarity with altered levels of the above components. CONCLUSIONS: The E-CELL system allows us to perform virtual experiments on the bacterial osmoregulation model. This model does not take into account interaction with other networks in the cell. It suggests that the regulation of OmpF and OmpC is a direct consequence of the level of OmpRP in the cell and is dependent on the way in which OmpRP interacts with ompF and ompC regulatory regions. The preliminary simulation experiment indicates that both reaching steady state expression and saturation is delayed in the case of OmpC compared to OmpF. Experimental analysis will help improve the model. The model captures the basic features of the generally accepted view of EnvZ-OmpR signaling and is a reasonable starting point for building sophisticated models and explaining quantitative features of the system.

Computer Simulation↗

PubMed2004

Analysis of the lambdoid prophage element e14 in the E. coli K-12 genome.

BACKGROUND: Many sequenced bacterial genomes harbor phage-like elements or cryptic prophages. These elements have been implicated in pathogenesis, serotype conversion and phage immunity. The e14 element is a defective lambdoid prophage element present at 25 min in the E. coli K-12 genome. This prophage encodes important functional genes such as lit (T4 exclusion), mcrA (modified cytosine restriction activity) and pin (recombinase). RESULTS: Bioinformatic analysis of the e14 prophage sequence shows the modular nature of the e14 element which shares a large part of its sequence with the Shigella flexneri phage SfV. Based on this similarity, the regulatory region including the repressor and Cro proteins and their binding sites were identified. The protein product of b1149 was found to be a fusion of a replication protein and a terminase. The genes b1143, b1151 and b1152 were identified as putative pseudogenes. A number of duplications of the stfE tail fibre gene of the e14 are seen in plasmid p15B. A protein based comparative approach using the COG database as a starting point helped detect lambdoid prophage like elements in a representative set of completely sequenced genomes. CONCLUSIONS: The e14 element was characterized for the function of its encoded genes, the regulatory regions, replication origin and homology with other phage and bacterial sequences. Comparative analysis at nucleotide and protein levels suggest that a number of important phage related functions are missing in the e14 genome including parts of the early left operon, early right operon and late operon. The loss of these genes is the result of at least three major deletions that have occurred on e14 since its integration. A comparative protein level approach using the COG database can be effectively used to detect defective lambdoid prophage like elements in bacterial genomes.

Bacillus subtilis↗

PubMed2004

HNH family subclassification leads to identification of commonality in the His-Me endonuclease superfamily.

The HNHc (SMART ID: SM00507) domain (SCOP nomenclature: HNH family) can be subclassified into at least eight subsets by iterative refinement of HMM profiles. An initial clustering of 323 proteins containing the HNHc domain helped identify the subsets. The subsets could be differentiated on the basis of the pattern of occurrence of seven defining features. Domain association is also different between the subsets. The subsets show organism as well as domain-based clustering, suggestive of propagation by both duplication and horizontal transfer events. Structure-based sequence analysis of the subsets led to the identification of common structural and sequence motifs in the HNH family with the other three families under the His-Me endonuclease superfamily.

Amino Acid Sequence↗

PubMed2004

A review of genetic and structural understanding of the role of myocilin in primary open angle glaucoma.

Primary open angle glaucoma (POAG) is the most common form of glaucoma and the second leading cause of blindness in the world. Discovery of the candidate gene MYOC (TIGR/MYOC) encoding the protein myocilin, believed to have a role in cytoskeletal function, might play a key role in understanding the pathogenesis of POAG. MYOC is expressed in many ocular tissues, including trabecular meshwork (TM), a specialised eye tissue essential in regulating intraocular pressure (IOP). Later it was shown to be the trabecular meshwork inducible-glucocorticoid response protein (TIGR). Mutations in MYOC have been identified as the cause of hereditary juvenile-onset open-angle glaucoma (JOAG). The unprocessed myocilin with signal peptide is a 55-kDa protein with 504 amino acids. Mature myocilin is known to form multimers. Wild type myocilin protein is normally secreted into the trabecular extracellular matrix (ECM) and there appears to interact with various ECM materials. It is believed that the deposition of high amounts of myocilin in trabecular ECM could affect aqueous outflow either by physical barrier and/or through cell-mediated process leading to elevation of IOP. The N-terminal region of the myocilin has sequence similarity to myosin (muscle protein) and the C-terminal of the protein has an olfactomedin-like domain. Structural and genetic studies of the MYOC gene and its protein product along with molecular modeling could lead to better understanding of the pathogenesis of POAG. This review highlights the current understanding of myocilin and the relevance of genetic and structural work.

Cytoskeletal Proteins↗

PubMed2004

Consensus based validation of membrane porins.

Porins are outermembrane beta-barrel proteins. They have varied biological functionality ranging from phage receptors, immunogenicity, pathogenicity to apoptosis. However, only a small number has been structurally and functionally characterised. A validation mechanism and a database of porins would be useful for target selection in proteomics and structural genomics work. Here we report a validation mechanism developed for membrane porins. A database server for porins, PRNDS, has been created containing experimentally proven porins and likely putative porins. Each porin is validated and ranked using a weighted scoring system developed based on six, structure and sequence based criteria. The server also predicts possible porins.

Amino Acid Sequence↗

PubMed2003

Evaluation and understanding of myocilin mutations in Indian primary open angle glaucoma patients.

PURPOSE: To screen for mutations in the MYOC gene of patients with Primary Open Angle Glaucoma (POAG) in India and to better understand the mutations using a possible model of myocilin. METHODS: We analyzed DNA for mutations in 107 subjects with POAG and 90 normal control subjects. The exonic sequences of the MYOC gene from all subjects were amplified by Polymerase Chain Reaction (PCR). We carried out Single Strand Conformation Polymorphism (SSCP) for all the PCR products. The DNA samples which showed mobility shift in the banding pattern in SSCP gel were sequenced. We also analyzed the presence of the common mutation Gln368Stop using a specific restriction enzyme Taa 1. The mutations observed here and elsewhere have been mapped onto a possible model built for myocilin using a knowledge-based consensus modeling approach. RESULTS: Two heterozygous mutations Gly367Arg (1099G>A) and Thr377Met (1130C>T) were identified in exon3 of the MYOC gene of probands 40-1 and 51-1 respectively, from material obtained from the 107 unrelated subjects with POAG. These two mutations were not present in the normal controls studied. We identified a Single Nucleotide Polymorphism (SNP) Gly122Gly (366C>T) in exon1 of proband 57-1 as a non-disease causing variation. The common mutation Gln368Stop found in the Western population was not observed in the POAG cases screened in Indian population. The possible structural model for myocilin suggests a predominantly [beta]-strand rich C-terminal region (181-504) which is connected by the [alpha]-helical mid-region (111-180) to the N-terminal region (34-110) which has low secondary structure content. Both the mutations, Gly367Arg and Thr377Met identified in our study, map on to the C-terminal region. These mutations disfavor burial of this region during oligomer formation due to the charged or bulky nature of the mutants. Most of the other mutations known for myocilin also are surface exposed on the C-terminal region. CONCLUSIONS: Our findings indicate that the mutation frequency of the MYOC gene is 2% in the Indian population affected with POAG, which is not a well-studied ethnic group of the Asian continent. The variations identified in our study have been previously reported in the Western population. The nonsense mutation Gln368Stop was not observed in the present study and thereby suggests that it may not be a common disease-causing mutation in the Indian population. Amongst other Asian populations, studies in Japan also didn't report this nonsense mutation. The location of these mutations suggest that a plausible mode of action could be by disruption of dimer or oligomer formation by the C-terminal region allowing greater chances of nucleation of aggregation by the N-terminal region.

Adolescent↗