Indian Journal of Biochemistry 
& Biophysics

 

Total visitors: 1,743  since 19-10-05

 

ISSN : 0301-1208

CODEN : IJBBBQ  

VOLUME 42

NUMBER 5

OCTOBER 2005

 

 
CONTENTS

 

CDC27 protein is involved in radiation response in squamous cell cervix carcinoma

271

T Rajkumar*, G Gopal, G Selvaluxmi and K R Rajalekshmy

 

 

 

Crystal structure of a novel phospholipase A2 from crude venom of Indian cobra sub-species Naja naja sagittifera at 1.48 resolution

279

Rajendra K Singh, Talat Jabeen, Sujata Sharma, Punit Kaur, A Srinivasan and
Tej P Singh*

 

 

 

Purification and characterization of a-amylase from Bacillus amyloliquefaciens
NCIM 2829

287

Mithu De, Kali P Das and P K Chakrabartty*

 

 

 

Effect of exogenous sucrose on the enzymes of starch degradation and sucrose metabolism in cowpea (Vigna unguiculata L.) seedlings

295

Narinder Kaur*, Harpreet Kaur and Anil K Gupta

 

 

 

A study of thermal denaturation/renaturation in DNA using laser light scattering: A new approach

301

Ritesh Kumar Dubey and D N Tripathi*

 

 

 

Analysis of solid-phase allele-specific primer extension characteristics on biochip in combination with modified primers and PicoGreen staining method

308

Shengying Qin and Jianhua Liu*

 

 

 

QSAR analysis of 2,4-diaminopyrido[2,3-d]pyrimidines and 2,4-diaminopyrrolo[2,3-d]-pyrimidines as dihydrofolate reductase inhibitors

315

P Jain, L K Soni, A K Gupta and S G Kashkedikar*

 

 

 

Notes

 

Evaluation of hepatoprotective potential of propolis extract in carbontetrachloride induced liver injury in rats

321

Sangeeta Shukla*, Monika Bhadauria and Anjana Jadon

 

 

 

Thermodynamic study of magnesium ion binding to a-amylase

326

Ali Akbar Saboury*, Setareh Ghasemi and Mohammad Umar Dahot

 

 

 

Instructions to Authors

 

 

 

*Author for correspondence

 

 

 

 

 

 

          

 

 

 

 

 

 

            AUTHOR INDEX

 

 


 Bhadauria M 321

 Chakrabartty P K   287

 Dahot M U 326

 Das K P                287

 De M                287

 Dubey R K                 301

 Ghasemi S              326

 Gopal G                271

 Gupta A K                315

 Gupta Anil K                 295

 Jabeen T               279

 Jadon A               321

 Jain P               315

 Kashkedikar S G                315

 Kaur H               295

 Kaur N               295

 Kaur P              279

 Liu J              308

 Qin S              308

 Rajalekshmy R               271

 Rajkumar T              271

 Saboury A A               326

 Selvaluxmi G               271

 Sharma S              279

 Shukla S              321

 Singh R K              279

 Singh T P              279

 Soni L K              315

 Srinivasan A               279

 Tripathi D N                301


 

 

 

 

 

Papers

 

 

 

Indian Journal of Biochemistry & Biophysics 

Vol. 42, October 2005, pp. 271-278

 

CDC27 protein is involved in radiation response in squamous cell cervix carcinoma

 

T Rajkumar*, G Gopal, G Selvaluxmi and K R Rajalekshmy

 

In the present study, an attempt was made to identify genes involved in radiation response in cervix carcinoma. Differential display technique was used to study the expression profiles of tumour biopsy samples obtained from patients, responding and not responding to treatment. The samples were obtained prior to radiotherapy and subsequent to treatment with Tele-radiation at 10 Gray (Gy). One of the differentially expressed cDNAs, when sequenced was identified to be CDC27. Immuno-histochemical analysis of pre- and post-treated tumour samples from fifteen patients showed the down-regulation of expression of CDC27 protein in seven patients. Down-regulation was associated with poorer response to radiotherapy. Cervical cancer cell lines SiHa and C33A were irradiated and their nuclei were stained for expression of CDC27 and analyzed using fluorescent-activated cell sorting (FACS). Down-regulation of CDC27 protein in the irradiated SiHa cell line was associated with greater survival fraction, compared to the irradiated C33A cell line, which had only slight fall in the level of CDC27 protein. This is the first study to suggest a role for CDC27 in radiation response. However, a larger cohort is needed to further confirm the value of CDC27 protein as a predictive marker, for radiation response in cervix cancer.

Keywords: cdc27, cervix cancer, predictive markers, radiation response, differential display, prognosis

 

IPC Code: G01N33/574

E -mail: cancer_institute_wia@vsnl.com

 

 

Indian Journal of Biochemistry & Biophysics 

Vol. 42, October 2005, pp. 279-286

 

Crystal structure of a novel phospholipase A2 from crude venom of Indian cobra sub-species Naja naja sagittifera at 1.48 resolution

Rajendra K Singh, Talat Jabeen, Sujata Sharma, Punit Kaur, A Srinivasan and Tej P Singh*

 

Secretory phospholipase A2s (PLA2s), the structurally-homologous enzymes share a common qualitative catalytic site, but differ greatly in their pharmacological properties and toxicities. There has been a recognizable pattern of mutations in the primary sequence of PLA2s that alter their catalytic properties significantly. In the present study, the amino acid sequence and the three-dimensional structure of a new isoform of PLA2 from crude venom of Indian cobra sub-species Naja naja sagittifera (N.n.s.) has been determined by X-ray crystallography. The crystal structure has revealed several novel features of PLA2 folding and function. It contains 913 protein atoms and one each of Ca2+, phosphate and acetate ions with 142 solvent water molecules. A Ca2+ ion is present in the calcium-binding loop and forms a seven-fold coordination with a distorted pentagonal bipyramidal geometry. One of the coordination linkages is with the acetate ion, instead of the conserved water molecule. The presence of Lys at position 31 has a stabilizing effect on the loop Tyr 25-Cys 29 by interacting with carbonyl oxygen atoms of Tyr 25, Gly 26 and Cys 29. In turn, it lends stability to the Ca2+-binding loop as well. Another unique feature of the PLA2 structure is the formation of an intrastrand hydrogen bond, involving Og of Thr 73 and Oe2 of Glu 71, thus helping the b-wing to act as a sharp arrow for insertion into other molecules. Yet another important feature of this PLA2 pertains to the conformation of its C-terminal segment, which is stabilized by a unique hydrogen bond through carbonyl oxygen of Lys 116 and Nd2 of Asn 120. This structural feature may be useful in the molecular recognition of the PLA2 through C-terminal segment.

Keywords: Phospholipase A2, crystal structure, Ca2+-binding loop, Naja naja sagittifera, crude venom, protein sequence

IPC Code: A61K35/58

E-Mail: tps@aiims.aiims.ac.in

 

 

Indian Journal of Biochemistry & Biophysics 

Vol. 42, October 2005, pp. 287-294

 

 

Purification and characterization of a-amylase from
Bacillus amyloliquefaciens NCIM 2829

Mithu De, Kali P Das and P K Chakrabartty*

 

a-Amylase (EC 3.2.1.1) was purified to homogeneity (specific activity 58,000 mmole min-1 mg protein-1) from the culture filtrate of Bacillus amyloliquefaciens NCIM 2829. Its molecular mass was found to be 67.5 kDa. The activity of the enzyme increased by almost 50% in the presence of Co+2 ion. Hg2+ and Cu2+ acted as strong inhibitors of the enzyme. The tryptophan moities of the enzyme were fairly protected from the aqueous environment. However, the globular interior of the protein was somewhat loosely packed. The protein had nearly an equal amount of a-helical and b-sheet structure in dilute solution. In concentrated solution, its secondary structure had a higher proportion of b-sheet at the expense of some random coil structure. The protein showed a molten globule state at a low concentration of chaotropic agent. The denaturation profile of the protein showed no cooperativity. Co2+ enhanced the structural stability of the enzyme.

Keywords: a-Amylase, Bacillus amyloliquefaciens NCIM 2829, circular dichroism, fluorescence measurement,
FT-IR spectroscopy, secondary structure, guanidine hydrochloride, metal ions, denaturation.

IPC Code: C12N 9/28

E-mail: krishna@boseinst.ernet.in

 

 

 

Indian Journal of Biochemistry & Biophysics 

Vol. 42, October 2005, pp. 295-300

 

 

Effect of exogenous sucrose on the enzymes of starch degradation and sucrose metabolism in cowpea (Vigna unguiculata L.) seedlings

 

Narinder Kaur*, Harpreet Kaur and Anil K Gupta

 

Addition of exogenous sucrose and mannitol in the growth medium decreased the germination and growth of cowpea (Vigna unguiculata L. cv C-88) seedlings. The reduced seedling growth appeared to be due to the decreased acid invertase activity in growing parts of the seedlings. An exogenous supply of sucrose upregulated the sucrose phosphate synthase (SPS) activity in different parts of seedlings. Decreased amylase activity in cotyledons and the mobilization of starch from cotyledons to the growing axis was observed in the presence of exogenous sucrose and mannitol. High sucrose content observed in different tissues in the presence of exogenous sucrose and mannitol was possibly due to high SPS and low acid invertase activities and reduced conversion of sucrose to starch. It appears that exogenous sucrose acts mainly as an osmoticum, rather than a source of carbon for the growing seedlings.

Keywords: Cowpea seedlings, Vigna unguiculata L., abiotic stress, sucrose, mannitol, germination, growth, amylase, invertase, sucrose synthase, sucrose phosphate synthase

IPC Code: C12N9/32; C08L3/00

E-mail: nkaur@rediffmail.com

 

 

Indian Journal of Biochemistry & Biophysics 

Vol. 42, October 2005, pp. 301-307

 

 

A study of thermal denaturation/renaturation in DNA using laser light scattering: A new approach

 

Ritesh Kumar Dubey and D N Tripathi*

 

The thermal denaturation/renaturation processes in E. coli and eukaryotic DNAs have been studied using the laser light scattering (LS) technique. The differential scattering intensity curve has been utilized to determine the transition temperature Tm. The effect of solution pH on DNA thermal denaturation has been examined. It has been shown clearly that LS is an extremely sensitive method (more than the UV absorption method) and reveals even the subtler effects such as the pre-transition fluctuations, and that the DNA denaturation is prominently affected by pH. The dependence of melting temperature (Tm) on composition of DNA, number of base pairs and base sequences has also been investigated. It has been observed that depending upon its base sequence, the Tm decreases in the case of renatured DNAs. The results have been compared with the UV absorption studies.

Keywords: Laser light scattering, DNA, denaturation, renaturation, E. coli, eukaryotic DNA

IPC Code: C07H21/04

 

E-mail: dnt@bhu.ac.in; dnt_bhu@yahoo.com

 

 

Indian Journal of Biochemistry & Biophysics 

Vol. 42, October 2005, pp. 308-314

 

  

Analysis of solid-phase allele-specific primer extension characteristics on biochip in combination with modified primers and
PicoGreen staining method

 

Shengying Qin and Jianhua Liu*

 

DNA microarray technology offers potential for future high-throughput variation genotyping. Allele-specific primer extension procedure on microarray has been considered as an efficient method for single nucleotide polymorphism (SNP) genotyping. However, the high cost of the fluorescent-labeled dNTP used for signal detection in this method limits its application. In the present study, we evaluated the characteristics of solid-phase allele-specific primer extension, in terms of specificity and efficiency and demonstrated that compared to liquid-phase reaction, it requires lower annealing temperature, and higher template and Mg2+concentrations. The extension efficiency and specificity were though linked, behave diametric during the gradient change of template and Mg2+concentrations or annealing temperature. To obtain both optimal signal intensity and specificity, we introduced an artificial mismatched base at the third position from the primer 3′end, which enhanced the specificity significantly. The PicoGreen staining method, which could decrease the cost greatly, was then introduced to replace the fluorescent-labeled dNTP for signal detection.

Keywords: Single nucleotide polymorphism, allele-specific primer extension, DNA microarray, extension efficiency and specificity, genotyping, PicoGreen staining

IPC Code: C07H21/04; C12N15/00; C1215/90

 

E-mail: Jianhualiudl@sjtu.edu.cn

 

 

 

Indian Journal of Biochemistry & Biophysics 

Vol. 42, October 2005, pp. 315-320

 

 

QSAR analysis of 2,4-diaminopyrido[2,3-d]pyrimidines and 2,4-diaminopyrrolo[2,3-d]pyrimidines as dihydrofolate reductase inhibitors

P Jain, L K Soni, A K Gupta and S G Kashkedikar*

 

Dihydrofolate reductase (DHFR) plays a ubiquitous role in the biosynthesis of DNA, RNA and essential amino acid methionine, and exhibits potential application in the treatment and prophylaxis of AIDS-associated opportunistic microbial infections. In this study, a series of DHFR analogs of 2,4-diaminopyrido[2,3-d]pyrimidines and 2,4-diaminopyrrolo[2,3-d]pyrimidines were subjected to quantitative structure-activity relationship (QSAR) analysis. The results showed that the electronic properties, energy of lowest unoccupied molecular orbital (LUMO) and Z-component of dipole moment (DPL3) of the molecule could be explored to design the potent DHFR inhibitors. LUMO is indicative of π-bonding interaction of species crucial for the electrophilicity of the molecules. This suggests that molecules are able to interact with electron-rich area at the receptor site. DPL3 is related to the molecular charge distribution in Z-component. These electronic parameters can be altered through the incorporation of electronegative groups. The QSAR study provides important structural insights for designing the potent DHFR inhibitors.

Keywords: dihydrofolate reductase, quantitative structure-activity relationship analysis, 2,4-diaminopyrido [2,3-d] pyrimidine, 2,4-diaminopyrrolo [2,3-d] pyrimidine, lowest unoccupied molecular orbital (LUMO), dipole moment

IPC Code: C12Q1/00,C12Q1/68

E-mail: arunkg_73@hotmail.com

 

 

 

Indian Journal of Biochemisty & Biophysics 

Vol. 42, October 2005, pp. 321-325

  

Notes

 

 

Evaluation of hepatoprotective potential of propolis extract in carbon tetrachloride induced liver injury in rats

Sangeeta Shukla*, Monika Bhadauria and Anjana Jadon

 

Propolis (bee glue), a resinous wax-like beehive product has been used since ancient times for its pharmaceutical properties. In the present study, the ethanolic extract of propolis (50, 100, 200 and 400 mg/kg, p.o.) was studied for its hepatoprotective activity against carbon tetrachloride (CCl4, 1.5 ml/kg, i.p.) induced liver damage in rats. Administration of CCl4 caused a sharp elevation in the activity of serum transaminases, serum alkaline phosphatase, acid phosphatase and hepatic lipid peroxidation (LPO) levels, and a significant decrease in the ATPase, alkaline phosphatase and succinic dehydrogenase activities in the liver and kidney and hepatic GSH level. The treatment with propolis extract at the doses of 200 and 400 mg/kg significantly reversed the various biochemical alterations in blood, liver and kidney induced by CCl4 intoxication. The hepatoprotective property of propolis may be due to its antioxidant activity.

Keywords: Propolis, hepatoprotective activity, rat, carbon tetrachloride, oxidative stress, liver function

IPC Code: A61P1/16; C11D3/16

 

E-mail: dr_sshukla@hotmail.com; monikabhadauria@rediffmail.com

 

 

 

Indian Journal of Biochemistry & Biophysics 

Vol. 42, October 2005, pp. 326-329

 

 

Thermodynamic study of magnesium ion binding to a-amylase

 

Ali Akbar Saboury*, Setareh Ghasemi and Mohammad Umar Dahot

The interaction of a-amylase (from Bacillus amyloliquefaciens) with Mg2+ ion was studied using UV spectrophotometric and isothermal titration calorimetric (ITC) methods at 27C in 30 mM Tris buffer solution at pH = 7.0. The binding isotherm for metal-protein interaction was easily obtained by carrying out ITC experiment at two different concentrations (2 mM and 50 mM) of the protein. a-Amylase had eight identical and independent binding sites for Mg2+ ion, which showed non-cooperativity in the binding process. The binding of Mg2+ ion was exothermic (DH= -17.3 kJ mol-1) with association binding constant of 2.08 mM-1. The binding slightly destabilized the enzyme against thermal denaturation, as evident from absorption studies.

Keywords: a-Amylase, magnesium ion, titration calorimetry, calorimetric method, metal binding, metal-protein interaction

IPC Code: C12N9/28

E-mail: saboury@ut.ac.ir