Cardiotoxin III from the Taiwan Cobra (Naja naja atra)

Bhaskaran, R.; Huang, Chung Chih; Chang, Ding Kwo; Yu, Chin

doi:10.1006/jmbi.1994.1082

Cited by 66 publications

(49 citation statements)

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“…4A) has a conformation that closely resembles that of the molecule in solution determined by 1 H NMR (yellow in Fig. 4A) (11,30). This suggests that L2 undergoes a local and specific conformational change in the presence of SDS.…”

Section: Resultsmentioning

confidence: 61%

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Structural Basis of Membrane-induced Cardiotoxin A3 Oligomerization

Forouhar

Huang

Liu

et al. 2003

Journal of Biological Chemistry

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Cobra cardiotoxins (CTXs) have previously been shown to induce membrane fusion of vesicles formed by phospholipids such as cardiolipin or sphingomyelin. CTX can also form a pore in membrane bilayers containing a anionic lipid such as phosphatidylserine or phosphatidylglycerol. Herein, we show that the interaction of CTX with negatively charged lipids causes CTX dimerization, an important intermediate for the eventual oligomerization of CTX during the CTX-induced fusion and pore formation process. The structural basis of the lipid-induced oligomerization of CTX A3, a major CTX from Naja atra, is then illustrated by the crystal structure of CTX A3 in complex with SDS; SDS likely mimics anionic lipids of the membrane under micelle conditions at 1.9-Å resolution. The crystal packing reveals distinct SDS-free and SDS-rich regions; in the latter two types of interconnecting CTX A3 dimers, D1 and D2, and several SDS molecules can be identified to stabilize D1 and D2 by simultaneously interacting with residues at each dimer interface. When the three CTX-SDS complexes in the asymmetric unit are overlaid, the orientation of CTX A3 monomers relative to the SDS molecules in the crystal is strikingly similar to that of the toxin with respect to model membranes as determined by NMR and Fourier transform infrared methods. These results not only illustrate how lipid-induced CTX dimer formation may be transformed into oligomers either as inverted micelles of fusion intermediates or as membrane pore of anionic lipid bilayers but also underscore a potential role for SDS in x-ray diffraction study of protein-membrane interactions in the future.

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Section: Resultsmentioning

confidence: 61%

“…Because interfaces of biologically relevant dimers usually bury 1400 Å 2 (34,35), it is unlikely that D1 and D2 can form in solution, as evidenced by NMR studies showing the existence of only monomeric species (11,30). Conversely, both dimers are likely to form when either SDS molecules or model membranes are present in the solution.…”

Section: Resultsmentioning

confidence: 99%

Structural Basis of Membrane-induced Cardiotoxin A3 Oligomerization

Forouhar

Huang

Liu

et al. 2003

Journal of Biological Chemistry

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“…Tyrosine residues located at positions 11 and 22 are lodged in the double-and triple-stranded ␤-sheet segments, respectively. Tyr 22 is one of the most well conserved residue found in all the cardiotoxin isoforms (17)(18)(19). This residue (Tyr 22 ) is the locus of a prominent hydrophobic core that plays an important role in maintaining the structural integrity of the three-dimensional structure of cardiotoxins.…”

Section: Resultsmentioning

confidence: 99%

Binding of Nucleotide Triphosphates to Cardiotoxin Analogue II from the Taiwan Cobra Venom (Naja naja atra)

Jayaraman

Krishnaswamy

Kumar

et al. 1999

Journal of Biological Chemistry

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Snake venom cardiotoxins have been recently shown to block the enzymatic activity of phospholipid protein kinase and Na ؉ ,K ؉ -ATPase. To understand the molecular basis for the inhibitory effects of cardiotoxin on the action of these enzymes, the nucleotide triphosphate binding ability of cardiotoxin analogue II (CTX II) from the Taiwan cobra (Naja naja atra) venom is investigated using a variety of spectroscopic techniques such as fluorescence, circular dichroism, and two-dimensional NMR. CTX II is found to bind to all the four nucleotide triphosphates (ATP, UTP, GTP, and CTP) with similar affinity. Detailed studies of the binding of dATP to CTX II indicated that the toxin molecule is significantly stabilized in the presence of the nucleotide. Molecular modeling, based on the NOEs observed for the dATP⅐CTX II complex, reveals that dATP binds to the CTX II molecule at the groove enclosed between the Nand C-terminal ends of the toxin molecule. Based on the results obtained in the present study, a molecular mechanism to account for the inhibition of the enzymatic activity of the phospholipid-sensitive protein kinase and Na ؉ ,K ؉ -ATPase is also proposed.Snake venom cardiotoxins are small molecular mass (ϳ7 kDa), highly basic proteins cross-linked by four disulfide bridges (1-3). These toxins exhibit a wide variety of biological activities such as contraction of cardiac muscle cells, lysis of erythrocytes, and selective toxicity to certain types of tumor cells (4, 5). More recently, cardiotoxins have been demonstrated to selectively inhibit the action of certain key enzymes such as Na ϩ ,K ϩ -ATPase and phospholipid-sensitive protein kinase (2, 6 -8). Although the general molecular mechanism underlying the enzyme inhibitory action of cardiotoxins is still an enigma, it is contemplated that cardiotoxin could effectively block the enzymatic activities of Na ϩ ,K ϩ -ATPase and phospholipid-sensitive protein kinase by competitively binding (in the present study) to adenosine triphosphate (ATP), which is a key substrate for the functioning of these enzymes. However, to date, the evidence for the binding of a nucleotide triphosphate to cardiotoxin has not been reported.In the present study, for the first time, we demonstrate that the cardiotoxin analogue II (CTX II) 1 isolated from the Taiwan cobra venom (Naja naja atra) bind to nucleotide triphosphates. Herein, we also propose a molecular model of the CTX II⅐dATP complex and suggest a reasonable molecular mechanism to explain the reported inhibitory action of snake venom cardiotoxin on phospholipid-sensitive protein kinase and Na ϩ ,K ϩ -ATPase. MATERIALS AND METHODSCTX II was purified as described by Yang et al. (9). The concentration of the protein was estimated from its absorbance at 280 nm (⑀ 1 mM ϭ 4.08). The nucleotides were purchased from Sigma. The concentration of the nucleotides was estimated (10) from their extinction coefficient values (⑀ 259 ϭ 15,400 for AMP, ADP, ATP, and dATP; ⑀ 271 ϭ 9,000 for CTP; ⑀ 253 ϭ 13,700 for GTP; and ⑀ 260 ϭ 1,000 for UTP). Al...

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“…For obtaining better spectral resolution, the two-dimensional homonuclear NMR experiments on CTX III were re-run on a 600 MHz Bruker DMX NMR spectrometer. We found that some of the resonance assignments reported earlier (19) were erroneous. Some of these errors in resonance assignments were also pointed out by Chiang et al (20).…”

Section: Fractionation Of the Crude Venom: Fractionation Of The Crudementioning

confidence: 55%

“…Structure Calculation: The three-dimensional solution structure of CTX HI was solved earlier by Bhaskaran et al (19) based on resonance assignments in the two dimensional 1H NMR spectra obtained on a 500 MHz Bruker DMX NMR spectrometer. For obtaining better spectral resolution, the two-dimensional homonuclear NMR experiments on CTX III were re-run on a 600 MHz Bruker DMX NMR spectrometer.…”

Section: Fractionation Of the Crude Venom: Fractionation Of The Crudementioning

confidence: 99%

The role of acetic acid in the prevention of salt‐induced aggregation of snake venom cardiotoxins

et al. 1998

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Summary: Snake venom cardiotoxins (CTXs) exhibit a strong tendency to aggregate upon desalting and hence it is extremely difficult to prepare salt-free cardiotoxin(s). In the present study, we describe a new method for preparation of salt-free CTX based on dialysis against acetic acid. Based on experimental observation and the three dimensional solution structure of cardiotoxin analogue I]I from the Taiwan cobra (Naja naja atra), a molecular mechanism for the prevention of aggregation of cardiotoxins by acetic acid is discussed. In our opinion, the results obtained in the present study would pave way for elucidating the structural basis for the broad spectrum of biological activities exhibited by snake venom cardiotoxins.Snake venoms are a mixture of many different proteins of which the cardiotoxins (1-5) and the neurotoxins (6-8) are the most toxic. Cardiotoxins exhibit an wide array of biological properties such as depolarization and contraction of muscular cells, lysis of erythrocytes, selective killing of certain types of cancer cells and inhibition of protein kinase C activity. Due to the broad spectrum of biological properties exhibited by the snake venom cardiotoxins, extensive research is being carried out to understand the structure-function relationship in this class of toxins (9,10).Cardiotoxins are highly basic (PI>I 0.0), small molecular weight (6.5-7 Kda), all [3-sheet proteins cross linked by four disulfide bridges (11-16). The isolation of cardiotoxins involves fractionation on a cation exchanger using a linear salt gradient (17).Owing to the highly positive nature of cardiotoxins, they elute only at high salt concentrations (> 0.2 M) (4). Presence of salt is known to affect the function of snake venom cardiotoxins significantly (18). However, interestingly,

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Cardiotoxin III from the Taiwan Cobra (Naja naja atra)

Cited by 66 publications

References 0 publications

Structural Basis of Membrane-induced Cardiotoxin A3 Oligomerization

Structural Basis of Membrane-induced Cardiotoxin A3 Oligomerization

Binding of Nucleotide Triphosphates to Cardiotoxin Analogue II from the Taiwan Cobra Venom (Naja naja atra)

The role of acetic acid in the prevention of salt‐induced aggregation of snake venom cardiotoxins

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