Synthesis and Characterization of the First Potent Inhibitor of Yapsin 1

Cawley, Niamh X.; Chino, Masao; Maldonado, Alex; Rodriguez, Yazmin M.; Loh, Y. Peng; Ellman, Jonathan A.

doi:10.1074/jbc.m207230200

Cited by 12 publications

(10 citation statements)

References 32 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…Pepstatin A was previously shown to be a potent inhibitor of secreted Sap activity (31,59) and of Sap9 activity at acidic pH (10). In our experiments, however, pepstatin A concentrations that inhibited Sap2 and Sap6 activity efficiently, were too low to elicit full Sap9 and Sap10 inhibition.…”

Section: Discussioncontrasting

confidence: 73%

Proteolytic Cleavage of Covalently Linked Cell Wall Proteins by Candida albicans Sap9 and Sap10

et al. 2011

View full text Add to dashboard Cite

The cell wall of the human-pathogenic fungus Candida albicans is a robust but also dynamic structure which mediates adaptation to changing environmental conditions during infection. Sap9 and Sap10 are cell surfaceassociated proteases which function in C. albicans cell wall integrity and interaction with human epithelial cells and neutrophils. In this study, we have analyzed the enzymatic properties of Sap9 and Sap10 and investigated whether these proteases cleave proteins on the fungal cell surface. We show that Sap9 and Sap10, in contrast to other aspartic proteases, exhibit a near-neutral pH optimum of proteolytic activity and prefer the processing of peptides containing basic or dibasic residues. However, both proteases also cleaved at nonbasic sites, and not all tested peptides with dibasic residues were processed. By digesting isolated cell walls with Sap9 or Sap10, we identified the covalently linked cell wall proteins (CWPs) Cht2, Ywp1, Als2, Rhd3, Rbt5, Ecm33, and Pga4 as in vitro protease substrates. Proteolytic cleavage of the chitinase Cht2 and the glucan-cross-linking protein Pir1 by Sap9 was verified using hemagglutinin (HA) epitope-tagged versions of both proteins. Deletion of the SAP9 and SAP10 genes resulted in a reduction of cell-associated chitinase activity similar to that upon deletion of CHT2, suggesting a direct influence of Sap9 and Sap10 on Cht2 function. In contrast, cell surface changes elicited by SAP9 and SAP10 deletion had no major impact on the phagocytosis and killing of C. albicans by human macrophages. We propose that Sap9 and Sap10 influence distinct cell wall functions by proteolytic cleavage of covalently linked cell wall proteins.

show abstract

Section: Discussioncontrasting

confidence: 73%

Proteolytic Cleavage of Covalently Linked Cell Wall Proteins by Candida albicans Sap9 and Sap10

et al. 2011

View full text Add to dashboard Cite

show abstract

“…Interestingly, the peptide KIHNKLFGF, which is identical to an internal sequence of Sap9 (Lys 149 to Phe 157 from the N terminus), was processed by Sap9 between Lys and Leu 154 . This suggests self-processing activity of Sap9 at this site, which possibly accounts for the two subunits observed in this study and by others (11). The N-terminal sequence of the larger ␤-subunit was previously shown to be Leu 154 -Phe-Gly-Phe (11), identical to the N-terminal sequence of one fragment of the digested peptide.…”

Section: Sap9 and Sap10 Are N-glycosylated And Gpi-anchored On The Cellsupporting

confidence: 82%

Glycosylphosphatidylinositol-anchored Proteases of Candida albicans Target Proteins Necessary for Both Cellular Processes and Host-Pathogen Interactions

Albrecht

Felk

Pichová

et al. 2006

Journal of Biological Chemistry

237

248

View full text Add to dashboard Cite

Intracellular and secreted proteases fulfill multiple functions in microorganisms. In pathogenic microorganisms extracellular proteases may be adapted to interactions with host cells. Here we describe two cell surface-associated aspartic proteases, Sap9 and Sap10, which have structural similarities to yapsins of Saccharomyces cerevisiae and are produced by the human pathogenic yeast Candida albicans. Sap9 and Sap10 are glycosylphosphatidylinositol-anchored and located in the cell membrane or the cell wall. Both proteases are glycosylated, cleave at dibasic or basic processing sites similar to yapsins and Kex2-like proteases, and have functions in cell surface integrity and cell separation during budding. Overexpression of SAP9 in mutants lacking KEX2 or SAP10, or of SAP10 in mutants lacking KEX2 or SAP9, only partially restored these phenotypes, suggesting distinct target proteins of fungal origin for each of the three proteases. In addition, deletion of SAP9 and SAP10 modified the adhesion properties of C. albicans to epithelial cells and caused attenuated epithelial cell damage during experimental oral infection suggesting a unique role for these proteases in both cellular processes and host-pathogen interactions.Proteases possess multiple functions in nature ranging from regulation of subtle cellular processes by activating distinct proproteins to nonspecific degradation of proteins for recycling of biomolecules. Several pathogenic microorganisms have adapted this biochemical property to fulfill a number of specialized functions during the infective process. The substrate specificities of these proteases may be very narrow, as in the case of bacterial toxins responsible for botulism or anthrax. In contrast, facultative pathogens may secrete proteases that have more general and much broader effects and play important roles in both saprophytic growth and infection.The yeast Saccharomyces cerevisiae has served as a eukaryotic model organism to study functions of regulatory proteases including the Kex2 protein, a proprotein processing subtilisin-like serine protease of the late Golgi compartment (1-5). Soon after the discovery of Kex2, numerous Kex2-like processing systems were identified in other eukaryotic species, including humans (6), plants (7), the fission yeast Schizosacchoromyces pombe (8), and the plant pathogenic fungus Ustilago maydis (9). Characteristic of target proteins of Kex2-like proteases are N-terminal processing sites with dibasic amino acids, in particular Lys-Arg. Although Kex2 was shown to be the major protease for processing at dibasic residues, alternative pathways exist in S. cerevisiae (10). Gene products of YPS1 (yapsin 1) and YPS2 (yapsin 2), two closely related glycosylphosphatidylinositol (GPI) 2 -anchored aspartic proteases, were able to cleave the ␣-factor pheromone precursor (a well known Kex2 substrate) at Lys-Arg sites (10 -12), and overexpression of YPS1 and YPS2 partially compensated for the loss of Kex2 (10, 13). However, the biological roles of yapsins are unknown.The human...

show abstract

“…Unlike Sap1-to -8p in C. albicans, Sap9p is apparently not secreted, and like YPS1, SAP9 expression is increased during stationary phase and in response to cell wall perturbation (11,43), adding support to the notion that YPS1 and SAP9 may have similar functions. Additionally, the sequence specificity of Yps1p and Sap9p appears to be reasonably similar because a peptide isostere-based inhibitor of Yps1p also has activity against Sap9p (9). Together, these considerations suggest that Sap9p may carry out similar functions in C. albicans.…”

Section: Discussionmentioning

confidence: 90%

Yapsins Are a Family of Aspartyl Proteases Required for Cell Wall Integrity in Saccharomyces cerevisiae

et al. 2005

View full text Add to dashboard Cite

The yeast cell wall is a crucial extracellular organelle that protects the cell from lysis during environmental stress and morphogenesis. Here, we demonstrate that the yapsin family of five glycosylphosphatidylinositollinked aspartyl proteases is required for cell wall integrity in Saccharomyces cerevisiae. Yapsin null mutants show hypersensitivity to cell wall perturbation, and both the yps1⌬2⌬ mutant and the quintuple yapsin mutant (5yps⌬) undergo osmoremedial cell lysis at 37°C. The cell walls of both 5yps⌬ and yps1⌬2⌬ mutants have decreased amounts of 1,3-and 1,6-␤-glucan. Although there is decreased incorporation of both 1,3-and 1,6-␤-glucan in the 5yps⌬ mutant in vivo, in vitro specific activity of both 1,3-and 1,6-␤-glucan synthesis is similar to wild type, indicating that the yapsins affect processes downstream of glucan synthesis and that the yapsins may be involved in the incorporation or retention of cell wall glucan. Presumably as a response to the significant alterations in cell wall composition, the cell wall integrity mitogen-activated kinase signaling cascade (PKC1-MPK pathway) is basally active in 5yps⌬. YPS1 expression is induced during cell wall stress and remodeling in a PKC1-MPK1-dependent manner, indicating that Yps1p is a direct, and important, output of the cell wall integrity response. The Candida albicans (SAP9) and Candida glabrata (CgYPS1) homologues of YPS1 complement the phenotypes of the yps1⌬ mutant. Taken together, these data indicate that the yapsins play an important role in glucan homeostasis in S. cerevisiae and that yapsin homologues may play a similar role in the pathogenic yeasts C. albicans and C. glabrata.The yapsins are a family of five glycosylphosphatidylinositol (GPI)-linked aspartyl proteases (YPS1 to -3, -6, and -7) in Saccharomyces cerevisiae that have homologues in other fungi such as Candida albicans (43), Candida glabrata (16), and Aspergillus oryzae (36). The founding members of this family, YPS1 and -2 (17, 32), were identified as suppressors of null mutations in KEX2, a trans-Golgi network-localized serine protease that processes secretory proteins such as pro-␣-factor and the exoglucanase Exg1p (3, 53). The remaining three yapsins were subsequently identified by sequence homology following completion of the S. cerevisiae genome sequence (47). Like Kex2p, three of the yapsins (Yps1p, -2p, and -3p) cleave proteins and peptides C terminal to basic residues, both in vivo and in vitro (8, 33). Although Kex2p is exquisitely selective for Lys-Arg pairs (53), the yapsins cleave C terminal to monobasic sites containing either Lys or Arg (33, 48). In vivo, Yps1p cleaves human mammalian prohormones such as ACTH and CCK (8) and both Yps1p and Yps2p display secretase-like activity toward ␤-amyloid precursor protein that is expressed in yeast (34,66). To date, however, no yeast substrates have been identified; moreover, the physiologic function of the yapsins in S. cerevisiae has been unclear.Previously reported work from this laboratory showed that the yapsin double nul...

show abstract

Synthesis and Characterization of the First Potent Inhibitor of Yapsin 1

Cited by 12 publications

References 32 publications

Proteolytic Cleavage of Covalently Linked Cell Wall Proteins by Candida albicans Sap9 and Sap10

Proteolytic Cleavage of Covalently Linked Cell Wall Proteins by Candida albicans Sap9 and Sap10

Glycosylphosphatidylinositol-anchored Proteases of Candida albicans Target Proteins Necessary for Both Cellular Processes and Host-Pathogen Interactions

Yapsins Are a Family of Aspartyl Proteases Required for Cell Wall Integrity in Saccharomyces cerevisiae

Contact Info

Product

Resources

About