Indian Journal of Biochemistry & Biophysics

CODEN : IJBBBQ    ISSN : 0301-1208

VOLUME 39

NUMBER 6

DECEMBER 2002

 
CONTENTS

 

Minireview

 

Dendritic cells as natural adjuvants and modulators of immune response in cancer immunotherapy

 

363

  Rajshekhar S Alli and Ashok Khar*

 

 

 

Papers

 

Effect of D-amino acids on the functional activity and conformational stability of

 

  ribonuclease-A

368

  Hanief M Shahjee, Vikas Rishi and Faizan Ahmad*

 

 

 

Activity of novel protein kinase C and distribution of protein kinase Cq  in subcellular fractions of normal and Duchenne muscular dystrophic muscle

 

377

  D V N Kumar, J Shanmugasundaram, C Sundaram  and M P J S Anandaraj*

 

Limited proteolysis of maize NADP-malic enzyme

382

  S Pinto, S R Rao and A S Bhagwat*

 

 

 

Matrix metalloprotease 2-mediated activation of Ca2+-ATPase by superoxide radical (O2·-) in plasma membrane of bovine pulmonary vascular smooth muscle

 

390

  Malay Mandal, Sudip Das, Tapati Chakraborti and Sajal Chakraborti*

 

Entrapment of Sorghum root oxalate oxidase into polyvinyl alcohol membrane

397

  Geeta Singh, Suman, Durgesh N Tanwar and C S Pundir*

 

 

 

Purification and characterization of lipoxygenase from aromatic and non-aromatic rice (Oryza sativa L.)

 

401

  Sanjay Basak and Rajendra Prasad Johari*
 

 

 

Transport through liquid membranes containing omeprazole and lansoprazole

406

  A N Nagappa*, P V Pandi, P K Mishra, Rahul K Girish and I Shanmukh

 

 

 

Synthesis, conformation and vibrational dynamics of the peptide -Ser-Cys-Lys-Leu-Asp-Phe-, a fragment of apolipoprotein B

 

410

  Shinoo Srivastava, Seema Srivastava, Girish Chandra Melkani, Shyam Singh,   Vishwambhar Dayal Gupta* and Vijai Prakash Gupta

 

 

 

Notes

 

Dietary regulation of adenosine deaminase activity in stomach, small intestine and spleen of mice

 

419

  Indrani Ray and Ramesh Sharma*

 

 

 

Ornithine α-ketoglutarate modulates the levels of antioxidants and lipid peroxidation products in ammonium acetate treated rats

 

422

  K B Dakshayani, S Velvizhi and P Subramanian*

 

 

 

Annual Index

425

 

——————

*Author for correspondence

 

 

 

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 363-367

 

Dendritic cells as natural adjuvants and modulators of immune response in cancer immunotherapy

Rajshekhar S Alli and Ashok Khar*

Received 16 July 2002

 

Dendritic cells (DCs) are potent antigen presenting cells. Mature DCs activate antigen specific naïve T cells, B cells and NK cells. Under certain conditions, DCs even silence T cell immune responses in vivo, thus, modulating the immune response. This special function of DCs could be exploited in the treatment of cancer, autoimmune disorder and chronic viral infections.

 

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 368-376

 

Effect of D-amino acids on the functional activity and conformational stability of ribonuclease-A

Hanief M Shahjee, Vikas Rishi# and Faizan Ahmad*

Received 22 Aptil 2002; revised 2 July 2002

Using cytidine 2¢:3¢ cyclic monophosphate as a substrate, Km and kcat of ribonuclease-A in the presence of different concentrations of D-amino acids (Ala, Ser, Pro and Lys) and their L-isomers were measured at pH 6.0 and 25°C. These kinetic parameters remained unchanged in the presence and absence of D-and L-amino acids. This is the first experimental evidence showing that D-amino acids are compatible with the enzyme function. Values of Tm (midpoint of denaturation), DHm (enthalpy change at Tm) and DCp (constant-pressure heat capacity change) were also determined from the heat-induced denaturation curves of the protein, measured in the presence and absence of D- and L-isomers of an amino acid at four different pH values. It is shown for the first time that these thermodynamic parameters, within experimental errors, do not depend on the stereospecificity of an amino acid. Estimates of DGDo with the help of Gibbs-Helmoltz equation (DGDo = DHm (1–298.15/Tm) - DCp [(Tm – 298.15) + 298.15 ln (298.15/Tm)]) using known values of Tm, DHm and DCp suggested that D- and L-amino acids are compatible with protein stability, for DGDo remained unchanged in the presence of amino acids.

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 377-381

 

Activity of novel protein kinase C and distribution of protein kinase Cq in subcellular fractions of normal and Duchenne muscular dystrophic muscle

 

D V N Kumar1, J Shanmugasundaram1, C Sundaram2 and M P J S Anandaraj1*

Received 5 May 2002; revised 16 October 2002

Phospholipid-dependent, Ca2+-independent isoenzymes termed novel protein kinase C or nPKC, include PKC d, e, h, q and m. Status and role of nPKC and PKCq in Duchenne muscular dystrophic (DMD) condition is unknown. In the present study, we have shown that most of the nPKC isoforms are translocated to the membrane fraction of DMD tissue specimen. It is well established that translocation plays a key role in signal transduction by individual PKC isoforms. In our experiment, the increased association of nPKC isoform PKCq to membrane was further confirmed by Western blot. Increased expression of PKCq mRNA was identified by dot blot analysis. The above results suggest that, the alterations in nPKC location and increased expression of PKCq observed is a result of modification of PKC-mediated signal transduction and cell function.

 

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 382-389

 

Limited proteolysis of maize NADP-malic enzyme

S Pinto, S R Rao and A S Bhagwat*

Received 16 April 2002; revised 30 September 2002

 

The incubation of maize malic enzyme at 37°C with trypsin at a ratio of 150:1 of malic enzyme to trypsin caused rapid and complete inactivation of enzyme activity. The inactivation was caused by fairly specific cleavage of the enzyme monomer (62 kDa) into 40 kDa and 20 kDa fragments. The intensity of 40 kDa band increased with the time of treatment of enzyme with trypsin from 2 to 30 min. Substrates, especially NADP (25 µM) provided almost total protection against trypsin inactivation of the enzyme activity. The studies carried out with various other endoproteases indicated that endoprotease Lys-C was most effective in inactivating malic enzyme activity. The kinetic properties of the truncated enzyme have been studied. The Km value for malate in case of native and modified enzyme was found to be identical. Km NADP for the modified enzyme was slightly higher indicating that after proteolysis the enzyme affinity for NADP had decreased. Limited proteolysis with trypsin did not show any appreciable change in fluorescence properties of the modified enzyme. Binding of NADPH to the enzyme was not affected after modification.

 

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 390-396

 

Matrix metalloprotease 2-mediated activation of Ca2+-ATPase by superoxide
radical (O2.-) in plasma membrane of bovine pulmonary vascular smooth muscle

Malay Mandal, Sudip Das, Tapati Chakraborti and Sajal Chakraborti*

Received 13 December 2001; revised and accepted 6 August 2002

The role of the matrix metalloprotease-2 (MMP-2) in regulating Ca2+-ATPase activity in bovine pulmonary artery smooth muscle plasma membranes during treatment with the O2.- generating system, hypoxanthine (HPX) plus xanthine oxidase (XO) has been studied. The smooth muscle membranes possess matrix metalloprotease (MMP) activity in gelatin zymogram, having an apparent molecular mass of 72 kDa; the activity is inhibited by the tissue inhibitor of metalloprotease-2 (TIMP-2). Since both protease and MMP-2 have same molecular mass and are inhibited by TIMP-2, it may, therefore, be suggested that the protease is the MMP-2. Treatment of the smooth muscle membrane suspension with the O2.- generating system stimulates MMP-2 activity, as evidenced by an apparent increase in the intensity of the protease activity. O2.- also enhances [14C]-gelatin degradation and Ca2+-ATPase activity. The increase in MMP activity, assessed by [14C]-gelatin degradation and Ca2+-ATPase activity are inhibited upon pretreatment with superoxide dismutase (SOD). The O2.- triggered MMP and Ca2+-ATPase activities in the membrane are found to be inhibited by TIMP-2. The stimulation of the MMP and Ca2+-ATPase activities remain unaffected by the inhibitors of serine, thiol and cysteine groups of proteases such as phenylmethylsulfonylfluoride (PMSF), Bowman Birk inhibitor (BBI), chymostatin, N-ethylmaleimide, leupeptin, antipain and pepstatin. Adding pure bovine MMP-2 to the smooth muscle membrane suspension causes an increase in Ca2+-ATPase activity, but the pretreatment with TIMP-2 inhibits the increase in the enzyme activity.

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 397-400

 

Entrapment of Sorghum root oxalate oxidase into polyvinyl alcohol membrane

 

Geeta Singh, Suman, Durgesh N Tanwar and C S Pundir*

Received 6 May 2002; revised 17 October 2002

A Cl- and NO3- insensitive oxalate oxidase, purified from the roots of 10-day old seedlings of grain Sorghum has been immobilized on polyvinyl alcohol (PVA) membrane through entrapment with 96.07% retention of initial activity. The membrane bound enzyme showed an increase in optimum pH (from 5.0 to 6.5), time of incubation (from 5 to 10 min) and Km for oxalate (from 0.38 to 6.23 mM), but decrease in incubation tempera­ture for maximum activity (from 37 to 30ºC) and Vmax (from 70 nmol/min/ml to 9.7 nmol H2O2/min) and was unaffected by Cl- and NO3. The membrane bound enzyme lost 50% of its initial activity after 30 days of storage at room temperature. The use of membrane bound oxalate oxidase in determination of serum oxalate of urinary stone patients is demonstrated.

 

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 401-405

 

Purification and characterization of lipoxygenase from aromatic and

non-aromatic rice (Oryza sativa L.)

 

Sanjay Basak and Rajendra Prasad Johari*

Received 27 March 2002; revised 2 September 2002

Lipoxygenase (Lox) has been extensively purified from aromatic (Bas-370) and non-aromatic (Pusa-834) rice varieties. Crude isolates of Lox from the aromatic varieties (Bas-370 and PB-1) showed higher specific activity (4-fold) when compared to non-aromatic varieties (Pusa-677 and Pusa-834). The activity was optimum at pH 8.0 in all four varieties. Anionic PAGE of Lox from three days old seedlings revealed one extra band (Rm 0.48) in aromatic varieties, besides the presence of a major band (Rm 0.28) in all the four varieties. Elution profile of Lox from Bas-370 and Pusa-834 on DEAE-cellulose column showed three distinct peaks (L-1, L-2 and L-3), L-2 being the major fraction in both the varieties. SDS-PAGE of purified L-2 from Bas-370 showed a single band of molecular mass ~88 kDa.

 

 

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 406-409

 

Transport through liquid membranes containing omeprazole and lansoprazole

A N Nagappa1, *, P V Pandi1, P K Mishra2, Rahul K Girish2 and I Shanmukh2

Received 17 September 2001; revised 1 May 2002

Omeprazole and lansoprazole, the therapeutically important drugs belonging to proton pump inhibitor category are extensively used in the treatment of gastric ulcers. Transport through liquid membranes generated by these drugs in lecithin-cholesterol mixture in series with a supporting membrane has been studied. The data obtained show the formation of liquid membrane in series with the supporting membrane. Transport of cations, chloride and bicarbonate ions in the presence liquid membranes generated by omeprazole and lanzoprazole indicate the modification in the permeability of various permeants.

 

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 410-418

 

Synthesis, conformation and vibrational dynamics of the peptide -Ser-Cys-Lys-Leu-Asp-Phe-, a fragment of apolipoprotein B

 

Shinoo Srivastava, Seema Srivastava, Girish Chandra Melkani, Shyam Singh, Vishwambhar Dayal Gupta*

and Vijai Prakash Gupta

Received 31 January 2002; revised 4 July 2002

The collective normal modes of the hexapeptide -Ser-Cys-Lys-Leu-Asp-Phe-, a fragment of apolipoprotein B (apo B), have been obtained. They reflect the dynamic nature and are atleast partly responsible for energy input in autolytic activity. Further, on energetic considerations based on the measurements reported by Sim & Sim, it has been shown that of the two such fragments only one induces autolysis, while the other remains anchored to the coated pit.

 

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 419-421

 

Dietary regulation of adenosine deaminase activity in stomach, small intestine and spleen of mice

 

Indrani Ray and Ramesh Sharma*

Received 22 April 2002; revised 28 October 2002

Activity of adenosine deaminase (ADA) and its regulation by dietary restriction were studied in the stomach, small intestine and spleen of mice. ADA activity (U/mg protein) was highest in the stomach, followed by small intestine and spleen of mice on normal diet. The activity decreased significantly in the stomach (41%) and small intestine (45%) of 24 hr fasted mice, when compared to mice fed ad-libitum. However, ADA activity in spleen did not show any change by dietary intervention. Refeeding of fasted mice for 24 hr restored the activity of ADA in tissues. In addition, dietary restriction (alternate days of feeding for three months) had a cumulative effect, whereby ADA activity decreased significantly in the stomach (53% on the day of feeding and 60% on the day of fasting) and small intestine (50% and 54% on the day of feeding and fasting, respectively) without any change in activity in spleen. These findings indicate that dietary restriction reduces ADA activity in a tissue-specific manner. Long-term dietary restriction leads to a cumulative adaptation in lowering the ADA activity of GIT, but not in spleen.

 

Indian Journal of Biochemistry & Biophysics

Vol. 39, December 2002, pp. 422-424

 

Ornithine α-ketoglutarate modulates the
levels of antioxidants and lipid peroxidation products in ammonium acetate treated rats

K B Dakshayani, S Velvizhi and P Subramanian*

Received 20 March 2002; revised 3 September 2002

The effects of ornithine α-ketoglutarate (OKG) on ammonium acetate induced hepatotoxicity were studied in experimental rats. The levels of urea, non-protein nitrogen and thiobarbituric acid reactive substances were significantly increased in ammonium acetate treated rats; but these levels were significantly decreased in ammonium acetate-OKG treated rats. Similar patterns were observed in the levels of free fatty acids, triglycerides and phospholipids. Furthermore, non-enzymatic (reduced glutathione) and enzymatic (glutathione peroxidase, superoxide dismutase and catalase) antioxidants were significantly decreased in ammonium acetate treated rats, when compared with control and were significantly increased in ammonium acetate-OKG treated rats compared to ammonium acetate treatment alone.