Structure of an aliphatic amidase from<i>Geobacillus pallidus</i>RAPc8

Kimani, Serah; Agarkar, Vinod B.; Cowan, Don A.; Sayed, Muhammed; Sewell, B. Trevor

doi:10.1107/s090744490703836x

Cited by 31 publications

(39 citation statements)

References 46 publications

(90 reference statements)

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“…The conserved Glu142 in the catalytic site acts as a general base to catalyze the hydrolysis of this intermediate (Kimani et al 2007). This confirmed the conservation of the Glu, Lys, Cys catalytic triad across the nitrilase superfamily members and also supports the classification of these amidases in the nitrilase superfamily.…”

Section: Structural and Biochemical Properties Of Amidasesmentioning

confidence: 98%

Amidases: versatile enzymes in nature

Sharma

Bhalla

2009

Rev Environ Sci Biotechnol

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Amidases are ubiquitous enzymes and biological functions of these enzymes vary widely. In past five decades, they turned out to be an attractive tool in industries for the synthesis of wide variety of carboxylic acids, hydroxamic acids and hydrazide, which find applications in commodity chemicals synthesis, pharmaceuticals agrochemicals and waste water treatments etc. Their proteins structures revealed that aliphatic amidases share the typical a/b hydrolase fold (like nitrilase superfamily) and signature amidases are evolutionary related to aspartic proteinases. They hydrolyse wide variety of amides (short chain aliphatic amides, mid-chain amides, arylamides, a-aminoamides and a-hydroxyamides) and can be grouped on the basis of their catalytic site and preferred substrate. They resist denaturation at extreme of pH and temperature because of their strong and compact multimeric structures. Inhibition studies and three-dimensional analysis of the structures identified a Glu59, Lys134, Cys166 catalytic triad and follow ''Bi-bi Ping-Pong'' mechanism reaction for amide hydrolysis and acyl transferase reactions. Many recombinant amidases have been expressed in Escherichia coli as well as in Brevibacterium lactofermentum.

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Section: Structural and Biochemical Properties Of Amidasesmentioning

confidence: 98%

Amidases: versatile enzymes in nature

Sharma

Bhalla

2009

Rev Environ Sci Biotechnol

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“…In both NitFhit and NitN, the tyrosine hydroxyl group forms a hydrogen bond across the dimer interface. In the aliphatic amidase represented by the structure with PDB code 2plq, the corresponding aspartate, D167, is also involved in an interaction across the dimer interface, in this case the formation of a salt bridge with a lysine, K278 (22). This indicates a general principle in which the formation of the dimer interface and the active site are connected rather than the requirement of a specific property for the residue following the active-site cysteine.…”

Section: Resultsmentioning

confidence: 99%

“…In addition, NitN lacked the 35-amino-acid C-terminal sequence found in the aliphatic amidases of G. pallidus (22) and P. aeruginosa (3).…”

Section: Resultsmentioning

confidence: 99%

“…2). The greatest sequence identities were to the formamidase AmiF from Helicobacter pylori, 32.9% (PDB code 2dyu) (18); the amidases from Pseudomonas aeruginosa, 30.6% (PDB code 2uxy) (4), and Geobacillus pallidus RAPc8, 29.9% (PDB code 2plq) (22); and the hypothetical protein PH0642 from P. horikoshii, 24.4% (PDB code 1j31) (34). In addition, the predicted fold of NitN superimposed on the ␣␤␤␣ monomer fold typical of the nitrilase superfamily enzymes (Fig.…”

Section: Resultsmentioning

confidence: 99%

“…For example, the potential roles of the additional catalytic glutamate residue E142 in the G. pallidus aliphatic amidase (22,39) and the conserved catalytic residues (EKEC), which may dictate either amide-or nitrile-hydrolyzing activity, are all targets for mutagenic and coordinated crystallographic analyses.…”

Section: Resultsmentioning

confidence: 99%

See 2 more Smart Citations

Unique Aliphatic Amidase from a Psychrotrophic and Haloalkaliphilic Nesterenkonia Isolate

Nel

Tuffin

Sewell

et al. 2011

Appl Environ Microbiol

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Nesterenkonia strain AN1 was isolated from a screening program for nitrile-and amide-hydrolyzing microorganisms in Antarctic desert soil samples. Strain AN1 showed significant 16S rRNA sequence identity to known members of the genus. Like known Nesterenkonia species, strain AN1 was obligately alkaliphilic (optimum environmental pH, 9 to 10) and halotolerant (optimum environmental Na ؉ content, 0 to 15% [wt/vol]) but was also shown to be an obligate psychrophile with optimum growth at approximately 21°C. The partially sequenced genome of AN1 revealed an open reading frame (ORF) encoding a putative protein member of the nitrilase superfamily, referred to as NitN (264 amino acids). The protein crystallized readily as a dimer and the atomic structure of all but 10 amino acids of the protein was determined, confirming that the enzyme had an active site and a fold characteristic of the nitrilase superfamily. The protein was screened for activity against a variety of nitrile, carbamoyl, and amide substrates and was found to have only amidase activity. It had highest affinity for propionamide but demonstrated a low catalytic rate. NitN had maximal activity at 30°C and between pH 6.5 and 7.5, conditions which are outside the optimum growth range for the organism.

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Microbial Transformation of Nitriles to High-Value Acids or Amides

Chen

Zheng

et al. 2009

Biotechnology in China I

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Biotransformation of nitriles mediated by nitrile-amide converting enzymes has attracted considerable attention and developed tremendously in the recent years in China since it offers a valuable alternative to traditional chemical reaction which requires harsh conditions. As a result, an upsurge of these promising enzymes (including nitrile hydratase, nitrilase and amidase) has been taking place. This review aims at describing these enzymes in detail. A variety of microorganisms harboring nitrile-amide converting activities have been isolated and identified in China, some of which have already applied with moderate success. Currently, a wide range of high-value compounds such as aliphatic, alicyclic, aromatic and heterocyclic amides and their corresponding acids were provided by these nitrile-amide degrading organisms. Simultaneously, with the increasing demand of chiral substances, the enantioselectivity of the nitrilase superfamily is widely investigated and exploited in China, especially the bioconversion of optically active alpha-substituted phenylacetamides, acids and 2,2-dimethylcyclopropanecarboxamide and 2,2-dimethylcyclopropanecarboxylic acid by means of the catalysts exhibiting excellent stereoselectivity. Besides their synthetic value, the nitrile-amide converting enzymes also play an important role in environmental protection. In this context, cloning of the genes and expression of these enzymes are presented. In the near future in China, an increasing number of novel nitrile-amide converting organisms will be screened and their potential in the synthesis of useful acids and amides will be further exploited.

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Structure of an aliphatic amidase fromGeobacillus pallidusRAPc8

Cited by 31 publications

References 46 publications

Amidases: versatile enzymes in nature

Amidases: versatile enzymes in nature

Unique Aliphatic Amidase from a Psychrotrophic and Haloalkaliphilic Nesterenkonia Isolate

Microbial Transformation of Nitriles to High-Value Acids or Amides

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