TY - JOUR
T1 - Identification, Biochemical Characterization, and In Vivo Detection of a Zn-Metalloprotease with Collagenase Activity from Mannheimia haemolytica A2
AU - Ramírez-Rico, Gerardo
AU - Martinez-Castillo, Moises
AU - Ruiz-Mazón, Lucero
AU - Meneses-Romero, Erika Patricia
AU - Palacios, José Arturo Flores
AU - Díaz-Aparicio, Efrén
AU - Abascal, Erasmo Negrete
AU - de la Garza, Mireya
N1 - Publisher Copyright:
© 2024 by the authors.
PY - 2024/1
Y1 - 2024/1
N2 - Respiratory diseases in ruminants are a main cause of economic losses to farmers worldwide. Approximately 25% of ruminants experience at least one episode of respiratory disease during the first year of life. Mannheimia haemolytica is the main etiological bacterial agent in the ruminant respiratory disease complex. M. haemolytica can secrete several virulence factors, such as leukotoxin, lipopolysaccharide, and proteases, that can be targeted to treat infections. At present, little information has been reported on the secretion of M. haemolytica A2 proteases and their host protein targets. Here, we obtained evidence that M. haemolytica A2 proteases promote the degradation of hemoglobin, holo-lactoferrin, albumin, and fibrinogen. Additionally, we performed biochemical characterization for a specific 110 kDa Zn-dependent metalloprotease (110-Mh metalloprotease). This metalloprotease was purified through ion exchange chromatography and characterized using denaturing and chaotropic agents and through zymography assays. Furthermore, mass spectrometry identification and 3D modeling were performed. Then, antibodies against the 110 kDa-Mh metalloprotease were produced, which achieved great inhibition of proteolytic activity. Finally, the antibodies were used to perform immunohistochemical tests on postmortem lung samples from sheep with suggestive histology data of pneumonic mannheimiosis. Taken together, our results strongly suggest that the 110-Mh metalloprotease participates as a virulence mechanism that promotes damage to host tissues.
AB - Respiratory diseases in ruminants are a main cause of economic losses to farmers worldwide. Approximately 25% of ruminants experience at least one episode of respiratory disease during the first year of life. Mannheimia haemolytica is the main etiological bacterial agent in the ruminant respiratory disease complex. M. haemolytica can secrete several virulence factors, such as leukotoxin, lipopolysaccharide, and proteases, that can be targeted to treat infections. At present, little information has been reported on the secretion of M. haemolytica A2 proteases and their host protein targets. Here, we obtained evidence that M. haemolytica A2 proteases promote the degradation of hemoglobin, holo-lactoferrin, albumin, and fibrinogen. Additionally, we performed biochemical characterization for a specific 110 kDa Zn-dependent metalloprotease (110-Mh metalloprotease). This metalloprotease was purified through ion exchange chromatography and characterized using denaturing and chaotropic agents and through zymography assays. Furthermore, mass spectrometry identification and 3D modeling were performed. Then, antibodies against the 110 kDa-Mh metalloprotease were produced, which achieved great inhibition of proteolytic activity. Finally, the antibodies were used to perform immunohistochemical tests on postmortem lung samples from sheep with suggestive histology data of pneumonic mannheimiosis. Taken together, our results strongly suggest that the 110-Mh metalloprotease participates as a virulence mechanism that promotes damage to host tissues.
KW - M. haemolytica
KW - collagen
KW - metalloproteases
KW - zymograms
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U2 - 10.3390/ijms25021289
DO - 10.3390/ijms25021289
M3 - Article
C2 - 38279292
AN - SCOPUS:85183380230
SN - 1661-6596
VL - 25
JO - International journal of molecular sciences
JF - International journal of molecular sciences
IS - 2
M1 - 1289
ER -