Structural and Kinetic Determinants of Aldehyde Reduction by Aldose Reductase
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- 作者：Sanjay Srivastava ; Stanley J. Watowich ; J. Mark Petrash ; Satish K. Srivastava ; and Aruni Bhatnagar
- 刊名：Biochemistry
- 出版年：1999
- 出版时间：January 5, 1999
- 年：1999
- 卷：38
- 期：1
- 页码：42 - 54
- 全文大小：133K
- 年卷期：v.38,no.1(January 5, 1999)
- ISSN：1520-4995
文摘
Aldose reductase (AR) is a member of the aldo-keto reductase superfamily. Due to its abilityto catalyze the formation of sorbitol from glucose during hyperglycemic and hypertonic stress, the aldose-reducing property of AR has been accepted as its main physiological and pathological function. Nonetheless,AR is a poor catalyst for glucose reduction and displays active-site properties unexpected of a carbohydrate-binding protein. We, therefore, examined the catalytic properties of AR with a series of naturally occurringaldehydes, compatible in their hydrophobicity to the large apolar active site of the enzyme. Our resultsshow that recombinant human AR is an efficient catalyst for the reduction of medium- to long-chainunbranched saturated and unsaturated aldehydes. The enzyme displayed selective preference for saturatedaldehydes, such as hexanal, and unsaturated aldehydes, such as trans-2-octenal and nonenal as well astheir 4-hydroxy derivatives. Short-chain aldehydes such as propanal and acrolein were reduced lessefficiently. Branched derivatives of acrolein or its glutathione conjugate (GS-propanal) were, however,reduced with high efficiency. In the absence of NADPH, the 
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unsaturated aldehydes caused covalentmodification of the enzyme. On the basis of electrospray mass spectrometric analysis of the wild-typeand site-directed mutants of AR (in which the solvent exposed cysteines were individually replaced withserine), the site of modification was identified to be the active-site residue, Cys 298. The unsaturatedaldehydes, however, did not modify the enzyme bound to NADPH and did not inactivate the enzymeduring catalysis. Modeling studies indicate that the large hydrophobic active site of AR can accommodatea large number of aldehydes without changes in the structure of the binding site or movement of sidechains. High hydrophobicity due to long alkyl chains or apolar substituents appears to stabilize theinteraction of the aldehyde substrates with the enzyme. Apparently, such hydrophobic interactions providesubstrate selectivity and catalytic efficiency of the order achievable by hydrogen bonding. Since severalof the aldehydes reduced by AR are either environmental and pharmacological pollutants or products oflipid peroxidation, the present studies provide the basis of future investigations on the role of AR inregulating aldehyde metabolism particularly under pathological states associated with oxidative stressand/or aldehyde toxicity.
, unsaturated aldehydes caused covalentmodification of the enzyme. On the basis of electrospray mass spectrometric analysis of the wild-typeand site-directed mutants of AR (in which the solvent exposed cysteines were individually replaced withserine), the site of modification was identified to be the active-site residue, Cys 298. The unsaturatedaldehydes, however, did not modify the enzyme bound to NADPH and did not inactivate the enzymeduring catalysis. Modeling studies indicate that the large hydrophobic active site of AR can accommodatea large number of aldehydes without changes in the structure of the binding site or movement of sidechains. High hydrophobicity due to long alkyl chains or apolar substituents appears to stabilize theinteraction of the aldehyde substrates with the enzyme. Apparently, such hydrophobic interactions providesubstrate selectivity and catalytic efficiency of the order achievable by hydrogen bonding. Since severalof the aldehydes reduced by AR are either environmental and pharmacological pollutants or products oflipid peroxidation, the present studies provide the basis of future investigations on the role of AR inregulating aldehyde metabolism particularly under pathological states associated with oxidative stressand/or aldehyde toxicity.