Purification and Characterization of Calpains From Pig Epidermis and Their Action on Epidermal Keratin

Ando, Yoshinobu; Imamura, Sadao; Kannagi, Reiji; Murachi, Takashi

doi:10.1111/1523-1747.ep12462532

Cited by 9 publications

(6 citation statements)

References 15 publications

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“…The Ca2+ requirements for activation of these calpains I and II (Figure 5) were in perfect agreement with those reported for authentic calpain I from pig erythrocytes and calpain II from pig kidney (Kitahara et al, 1984). These results clearly indicate that the intact form of calpain I in PMN cells should have a subunit composition of 83 plus 29 kDa, as a heterodimer of the same composition has been found in many other tissues and cells of pigs (Kitahara et al, 1984;Yoshimura et al, 1984;Hatanaka et al, 1984;Ando et al, 1988). We thus concluded that the 70-kDa monomeric calpain I, calpain IM, found and isolated on nitrogen cavitation at 335 psi, is an artifact formed from heterodimeric calpain I, calpain ID, during the course of the cell disruption.…”

Section: Discussionsupporting

confidence: 64%

Molecular and catalytic characterization of intact heterodimeric and derived monomeric calpains isolated under different conditions from pig polymorphonuclear leukocytes

Fukui

Toyohara²,

Ito³

et al. 1988

Biochemistry

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Evidence is presented of polymorphonuclear (PMN) cells derived from pig peripheral blood containing two molecular species of Ca2+-dependent cysteine endopeptidases, calpains I and II, which require low and high concentrations of Ca2+, respectively, for activation. Calpains I and II, purified from PMN homogenates, are heterodimers consisting of 83 plus 29 kDa and 80 plus 29 kDa subunits, respectively, which can be identified by using subunit-specific antibodies and which are identical with those of calpain species in other pig tissues and cells hitherto reported. However, a 70-kDa calpain can also be detected when pig PMN cells are disrupted by the nitrogen cavitation method under rather mild conditions, i.e., with minimal destruction of the lysosomes. Lines of evidence are presented showing that the 70-kDa species is devoid of the light subunit, that it is artificially derived from naturally occurring heterodimeric calpain I, and that the PMN cells before disruption contained no such monomeric form. The isolated 70-kDa calpain I, or monomeric artifact, requires only 1 microM Ca2+ for half-maximal activation, and it is less pH stable and much less heat stable than the parent heterodimeric calpain I. A possible mechanism for the production of this artifact is discussed.

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Section: Discussionsupporting

confidence: 64%

Molecular and catalytic characterization of intact heterodimeric and derived monomeric calpains isolated under different conditions from pig polymorphonuclear leukocytes

Fukui

Toyohara²,

Ito³

et al. 1988

Biochemistry

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“…Our results (Figure 2A) showed a calcium-dependent effect on the generation of fragments from full-length K6A in human corneal epithelial cells, and UPP degradation of host proteins can be increased by elevated intracellular calcium (57). However, generation and degradation of KDAMPs could also reflect the activities of calcium-dependent proteases, e.g., calpains, which are known to target keratins (58,59) and are present in corneal epithelial cells and other ocular tissues (60). Further studies are underway to investigate the mechanisms of KDAMP generation and regulation in epithelial cells.…”

Section: Methodsmentioning

confidence: 72%

Cytokeratins mediate epithelial innate defense through their antimicrobial properties

Tam

Mun²,

Evans³

et al. 2012

J. Clin. Invest.

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Epithelial cells express antimicrobial proteins in response to invading pathogens, although little is known regarding epithelial defense mechanisms during healthy conditions. Here we report that epithelial cytokeratins have innate defense properties because they constitutively produce cytoprotective antimicrobial peptides. Glycine-rich C-terminal fragments derived from human cytokeratin 6A were identified in bactericidal lysate fractions of human corneal epithelial cells. Structural analysis revealed that these keratin-derived antimicrobial peptides (KDAMPs) exhibited coil structures with low α-helical content. Synthetic analogs of these KDAMPS showed rapid bactericidal activity against multiple pathogens and protected epithelial cells against bacterial virulence mechanisms, while a scrambled peptide showed no bactericidal activity. However, the bactericidal activity of a specific KDAMP was somewhat reduced by glycine-alanine substitutions. KDAMP activity involved bacterial binding and permeabilization, but the activity was unaffected by peptide charge or physiological salt concentration. Knockdown of cytokeratin 6A markedly reduced the bactericidal activity of epithelial cell lysates in vitro and increased the susceptibility of murine corneas to bacterial adherence in vivo. These data suggest that epithelial cytokeratins function as endogenous antimicrobial peptides in the host defense against infection and that keratin-derived antimicrobials may serve as effective therapeutic agents.

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“…Urocanic acid (UCA) and pyrrolidone carboxylic acid (PCA), are two transformed FLG breakdown products and are often used as markers for NMF and together with FAA have been used to assess SC moisturization in different dry skin types . Their production by the enzymatic processing of profilaggrin/FLG has been of recent interest with several groups examining the role of calpain‐1 (C‐1) and bleomycin hydrolase (BH) in the later stages of FLG degradation .…”

Section: Introductionmentioning

confidence: 99%

“…In contrast, C‐1 is a heterodimeric calcium‐activated cysteine protease comprised of a large subunit (80 kDa) and a small subunit (30 kDa). C‐1 is known to be present in human and porcine epidermis and has been reported to play an important role in the generation of NMF . It also has a direct role in epidermal differentiation by activation of epidermal transglutaminases .…”

Section: Introductionmentioning

confidence: 99%

Variation in the activities of late stage filaggrin processing enzymes, calpain‐1 and bleomycin hydrolase, together with pyrrolidone carboxylic acid levels, corneocyte phenotypes and plasmin activities in non‐sun‐exposed and sun‐exposed facial stratum corneum of different ethnicities

Raj

Voegeli

Rawlings

et al. 2016

Intern J of Cosmetic Sci

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Variation in PCA levels has been found for different ethnic groups in this study (AA > BA > C subjects). The values in the AA subjects are surprising as one might expect that the lack of pigmentation, and thereby increased photodamage, might lead to lower levels. Increased BH, but not C-1 activity, was observed in the AA subjects indicating that BH is associated with PCA production to a greater extent. Surprisingly, corneocyte maturation is still impaired with elevated PCA levels in AA subjects. The higher levels of plasmin and BH activities on the cheeks, especially for AA and C subjects, suggest that they can be used as markers for epidermal photodamage.

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Purification and Characterization of Calpains From Pig Epidermis and Their Action on Epidermal Keratin

Cited by 9 publications

References 15 publications

Molecular and catalytic characterization of intact heterodimeric and derived monomeric calpains isolated under different conditions from pig polymorphonuclear leukocytes

Molecular and catalytic characterization of intact heterodimeric and derived monomeric calpains isolated under different conditions from pig polymorphonuclear leukocytes

Cytokeratins mediate epithelial innate defense through their antimicrobial properties

Variation in the activities of late stage filaggrin processing enzymes, calpain‐1 and bleomycin hydrolase, together with pyrrolidone carboxylic acid levels, corneocyte phenotypes and plasmin activities in non‐sun‐exposed and sun‐exposed facial stratum corneum of different ethnicities

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