Aminopeptidase yscCo-II: a new cobalt-dependent aminopeptidase from yeast?purification and biochemical characterization

Herrera‐Camacho, Irma; Morales‐Monterrosas, Rosalva; Quiróz‐Alvarez, Rubén

doi:10.1002/(sici)1097-0061(200002)16:3<219::aid-yea523>3.0.co;2-j

Cited by 17 publications

(22 citation statements)

References 37 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The molecular mass of the purified enzyme was 46 kDa as determined by gel filtration and 40 kDa through SDS-PAGE, suggesting that the native enzyme is mo-nomeric. This contrasts with results obtained for other yeast aminopeptidases; in S. cerevisiae, the AP1 aminopeptidase has a molecular mass of 640 kDa, constituted by 12 subunits [26]; the APCo-II depicts a molecular mass of 290 kDa, constituted by 6 subunits [27]. In S. pombe, a dimeric 184 kDa aminopeptidase has been described [15].…”

Section: Discussioncontrasting

confidence: 75%

“…This inhibitory effect has been observed in other aminopeptidases isolated from yeast, such as AP-Y from S. cerevisiae [31] and aminopeptidase I from S. pombe [15]. Bestatin, an inhibitor of metalloenzymes, exerted a strong inhibitory effect on aminopeptidase from K. marxianus, similar to observed on AP-CoII aminopeptidase from S. cerevisiae [27]. However, this inhibitor has no effect on APCo-I [32], AP-yscII [27] and AP-Y [31] S. cerevisiae aminopeptidases.…”

Section: Discussionsupporting

confidence: 73%

“…Bestatin, an inhibitor of metalloenzymes, exerted a strong inhibitory effect on aminopeptidase from K. marxianus, similar to observed on AP-CoII aminopeptidase from S. cerevisiae [27]. However, this inhibitor has no effect on APCo-I [32], AP-yscII [27] and AP-Y [31] S. cerevisiae aminopeptidases. The presence of a blocking agent from thiol groups, such as E-64, inhibited partially the activity, suggesting that a thiol group might be participating at the catalytic site or near to it.…”

Section: Discussionmentioning

confidence: 77%

See 2 more Smart Citations

Purification and characterization of a lysine aminopeptidase from Kluyveromyces marxianus

Ramı́rez-Zavala

Mercado-Flores

Hernández‐Rodríguez

et al. 2004

FEMS Microbiology Letters

View full text Add to dashboard Cite

A lysine aminopeptidase was purified from the yeast Kluyveromyces marxianus. This enzyme was purified 100-fold from a soluble extract obtained at 100,000g. The purification procedure consisted in fractionated precipitation with ammonium sulfate and five chromatography steps. The native enzyme had a molecular mass of 46 kDa assessed through gel filtration. This aminopeptidase depicted an optimal pH of 7.0 and was stable at a pH range of 4-8, its optimal temperature was 45 degrees C and the enzyme became unstable at temperatures above 55 degrees C. The isoelectric point of the purified enzyme was 4.4. Michaelis constant and Vmax for L-lysine-p-nitroanilide were 0.33 mM and 2.2 mM min(-1) per milligram of protein, respectively. The enzyme was strongly inhibited by bestatin, o-phenanthroline and, to a lesser extent, by EDTA, suggesting that this enzyme is a metalloprotease. Our results suggest that the lysine aminopeptidase from Kluyveromyces marxianus might be of biotechnological relevance.

show abstract

Section: Discussioncontrasting

confidence: 75%

Section: Discussionsupporting

confidence: 73%

Section: Discussionmentioning

confidence: 77%

See 1 more Smart Citation

Purification and characterization of a lysine aminopeptidase from Kluyveromyces marxianus

Ramı́rez-Zavala

Mercado-Flores

Hernández‐Rodríguez

et al. 2004

FEMS Microbiology Letters

View full text Add to dashboard Cite

show abstract

“…Lysine aminopeptidases with substrate specificities similar to that of the P. furiosus enzyme have not been reported from any prokaryote; they have only been found in the fungi A. niger (6), S. cerevisiae (20), and K. marxianus (33). The S. cerevisiae enzyme, like that of P. furiosus, is also stimulated by Co 2ϩ ions (0.5 mM); however, the A. niger and K. marxianus enzymes do not require added metal ions.…”

Section: Discussionmentioning

confidence: 99%

“…However, these typically hydrolyze a broad range of peptides and are not specific for basic amino acids (25,32). On the other hand, three lysyl-specific aminopeptidases have been characterized from eukaryotic sources: the native form of a cobalt-dependent enzyme, yscCo-II, from the unicellular eukaryote Saccharomyces cerevisiae (20), the native enzyme from the yeast Kluyveromyces marxianus (33), and a recombinant form of a zinc-dependent enzyme from the filamentous fungus Aspergillus niger (6). The majority of aminopeptidases characterized so far, including the S. cerevisiae and A. niger enzymes, belong to the large M1 family of metallopeptidases.…”

mentioning

confidence: 99%

Characterization of a Novel Zinc-Containing, Lysine-Specific Aminopeptidase from the Hyperthermophilic ArchaeonPyrococcus furiosus

et al. 2005

View full text Add to dashboard Cite

Cell extracts of the proteolytic, hyperthermophilic archaeon Pyrococcus furiosus contain high specific activity (11 U/mg) of lysine aminopeptidase (KAP), as measured by the hydrolysis of L-lysyl-p-nitroanilide (Lys-pNA). The enzyme was purified by multistep chromatography. KAP is a homotetramer (38.2 kDa per subunit) and, as purified, contains 2.0 ؎ 0.48 zinc atoms per subunit. Surprisingly, its activity was stimulated fourfold by the addition of Co 2؉ ions (0.2 mM). Optimal KAP activity with Lys-pNA as the substrate occurred at pH 8.0 and a temperature of 100°C. The enzyme had a narrow substrate specificity with di-, tri-, and tetrapeptides, and it hydrolyzed only basic N-terminal residues at high rates. Mass spectroscopy analysis of the purified enzyme was used to identify, in the P. furiosus genome database, a gene (PF1861) that encodes a product corresponding to 346 amino acids. The recombinant protein containing a polyhistidine tag at the N terminus was produced in Escherichia coli and purified using affinity chromatography. Its properties, including molecular mass, metal ion dependence, and pH and temperature optima for catalysis, were indistinguishable from those of the native form, although the thermostability of the recombinant form was dramatically lower than that of the native enzyme (half-life of approximately 6 h at 100°C). Based on its amino acid sequence, KAP is part of the M18 family of peptidases and represents the first prokaryotic member of this family. KAP is also the first lysine-specific aminopeptidase to be purified from an archaeon.Aminopeptidases are exopeptidases that catalyze the removal of amino acid residues at the N termini of peptides and proteins. Intracellular proteolytic degradation by aminopeptidases is necessary for modulation of protein concentrations, maintenance of amino acid pools, and removal of damaged proteins (17). In addition, aminopeptidases have more specific functions, including activation (10) and inactivation (21) of biologically active peptides and the removal of N-terminal methionyl residues of newly synthesized proteins. The breakdown of proteinaceous substrates, whether imported into the cell or generated from damaged proteins, are further degraded by di-, tri-, and carboxypeptidases, as well as by aminopeptidases (17). Classification of aminopeptidases has been generally based upon their substrate specificities, such as preference for a neutral, acidic, or basic amino acid in the P1 position of the amino terminus of peptides (44). In recent years, more focus has been placed on classifying peptidases based on structural analyses (35). Aminopeptidases are widely distributed among eukaryotes and prokaryotes, although only a few have been isolated from archaea (37). About two-thirds of all aminopeptidases are represented by the metal-dependent peptidases in which zinc is the most frequently associated metal (34).Hyperthermophilic archaea are potentially rich sources of peptidase-type enzymes, because most of them are capable of using protein-based substrates as ...

show abstract

aminopeptidase S 3.4.11.24

Schomburg¹,

Schomburg²

2013

Class 3.4–6 Hydrolases, Lyases, Isomerases, Ligases

View full text Add to dashboard Cite

Aminopeptidase yscCo-II: a new cobalt-dependent aminopeptidase from yeast?purification and biochemical characterization

Cited by 17 publications

References 37 publications

Purification and characterization of a lysine aminopeptidase from Kluyveromyces marxianus

Purification and characterization of a lysine aminopeptidase from Kluyveromyces marxianus

Characterization of a Novel Zinc-Containing, Lysine-Specific Aminopeptidase from the Hyperthermophilic ArchaeonPyrococcus furiosus

aminopeptidase S 3.4.11.24

Contact Info

Product

Resources

About