In Silico Parameters of Methioninase of Pseudomonas Putida

Cancer might affect people at all ages, even fetuses , but the threat for most varieties increases with age (Merlo et al., 2006). Cancer causes about 13% of all deaths (Sasco et al., 2004).A methionine gamma-lyase (EC 4.4.1.11) is an enzyme that catalyzes the chemical reaction. Thus, the two substrates of this enzyme are L-methionine and H2O, whereas its 3 products are methanethiol, NH3, and 2-oxobutanoate. This enzyme belongs to the family of lyases, specifically the class of carbon-sulfur lyases. The systematic name of this enzyme class is L-methionine methanethiol-lyase (deaminating 2-oxobutanoate-forming). Other names in common use include L-methioninase, methionine lyase, methioninase, methionine dethiomethylase, L-methionine Ƴ lyase, and L-methionine methanethiol-lyase (deaminating). This enzyme participates in selenoamino acid metabolism. It employs one cofactor, pyridoxal phosphate (Kreis and Hession, 1973). Methionine dependence, the elevated methionine requirement for tumor cell proliferation, is the property of the majority of tumor cell types tested (Hoffman and Erbe 1976 and Hoffman et al., 1979). There have been several therapeutic strategies to target the methionine dependence of cancer cells. Methionine starvation therapy, such as with a methionine-free diet or methioninedepleted total parenteral nutrition, prolonged the survival time of tumor-bearing rodents (Goseki et al., 1992). Methionine-free total parenteral nutrition in combination with chemotherapeutic drugs extended the survival of patients with high-stage gastric cancer (Goseki et al., 1995). METase from Pseudomonas putida, which degrades extracellular methionine to a-ketobutyrate, ammonia, and methanethiol (Tanaka et al., 1977), has been demonstrated to have antitumor efficacy in vitro and in vivo (Tan et al., 1997 ). METase has synergistic efficacy in combination with 5-fluorouracil (Yoshioka et al., 1998) and cisplatin (Tan et al., 1999).

Numerous cell culture studies using normal and malignant cell lines (e.g., leukemia, prostate) demonstrated that methionine restriction suppresses cancer cell growth, with little or no deleterious effect on normal cells by Poirson-Bichat (1997). Likewise, tumors are methionine dependent in vivo. Dietary methionine restriction causes regression of a variety of animal tumors and inhibits metastasis in animal models reported by Guo et al., (1993) and Millis et al., (1998).

Methionine Ƴ lyase (EC 4.4.1.11) (L-methioninase) Pseudomonas putida protein parameters were analysed by using the tool Prosite at www.expasy.org/prosite. All the protein parameters with respect to amino acid composition, secondary structure prediction, hydrophobicity, isoelectric point etc were analyzed.

Methionine gamma-lyase (L-methioninase) Pseudomonas putida.The computation has been

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Number of amino acids: 398, Molecular weight: 42626.7, Theoretical pI: 6.21 Total number of negatively charged residues (Asp + Glu): 36 Total number of positively charged residues (Arg + Lys): 29 Extinction coefficients Extinction coefficients are in units of M-1 cm-1, at 280 nm measured in water.Ext. Coefficient 25120, Abs 0.1% (=1 g/l) 0.589, assuming ALL Cys residues appear as half cystines Estimated half-life The N-terminal of the sequence considered is M (Met).The estimated half-life is: 30 hours (mammalian reticulocytes, in vitro).>20 hours (yeast, in vivo).>10 hours (Escherichia coli, in vivo). Instability index The instability index (II) is computed to be 37.09. This classifies the protein as stable. Aliphatic index: 91.31.Grand average of hydropathicity (GRAVY): 0.026 Molecular weight (average): 42626.70, Theoretical pI: 6.21.

Methionine gamma-lyase (L-methioninase) of Pseudomonas putida. The parameters have been computed for the following feature FT CHAIN 1398 Methionine gamma-lyase. The computation has been carried out on the complete sequence (398 amino acids).

SEQUENCE LENGTH: 398

Fig.1b: Pepwheel of methioninase of Pseudomonas putida

Fig.2: SOPMA of methioninase of Pseudomonas putida

Main Ramachandran plot

The Ramachandran plot shows the phi-psi torsion angles for all residues in the structure (except those at the chain termini). Glycine residues are separately identified by triangles as these are not restricted to the regions of the plot appropriate to the other sidechain types. The colouring/shading on the plot represents the different regions (see below) described in Morris et al. (1992): the darkest areas (here shown in red) correspond to the "core" regions representing the most favourable combinations of phi-psi values. Ideally, one would hope to have over 90% of the residues in these "core" regions. The percentage of residues in the "core" regions is one of the better guides to stereochemical quality. The different regions on the Ramachandran plot are as described in Morris et al. (1992). The regions are labelled as follows:

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The different regions were taken from the observed phi-psi distribution for 121,870 residues from 463 known X-ray protein structures. The two most favoured regions are the "core" and "allowed" regions which correspond to 10° x 10° pixels having more than 100 and 8 residues in them, respectively. The "generous" regions were defined by Morris et al. (1992) by extending out by 20° (two pixels) all round the "allowed" regions. In fact, the authors found very few residues in these "generous" regions, so they can probably be treated much like the "disallowed" region and any residues in them investigated more closely.

The plot shows separate Ramachandran plots for each of the 20 different amino acid types. The darker the shaded area on each plot, the more favourable is the region. The data on which the shading is based has come from a data set of 163 non-homologous, high-resolution protein chains chosen from structures solved by X-ray crystallography to a resolution of 2.0Å or better and an R-factor no greater than 20%. The numbers in brackets, following each residue name, show the total number of data points on that graph. The red numbers above the data points are the reside-numbers of the residues in question (ie showing those residues lying in unfavourable regions of the plot).

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