Binding of monoclonal antibody to cathepsin M located on the external surface of rabbit lysosomes

Area of Science:

• Biochemistry
• Cell Biology
• Immunology

Background:

• Cathepsin M is a membrane-bound enzyme found in rabbit liver lysosomes.
• Previous research indicated that trypsin can digest membrane-bound cathepsin M.

Purpose of the Study:

• To confirm the location of membrane-bound cathepsin M on the lysosomal membrane.
• To investigate the specificity of antibody binding to cathepsin M.

Main Methods:

• Generation of a monoclonal antibody against rabbit liver cathepsin M.
• Incubation of intact rabbit liver lysosomes with the antibody.
• Treatment of lysosomes with trypsin to assess antibody binding.
• Affinity chromatography using Sepharose coupled to anti-cathepsin M antibody.

Main Results:

• The monoclonal antibody specifically binds to intact rabbit liver lysosomes.
• Trypsin treatment of lysosomes abolished the antibody binding, consistent with cathepsin M digestion.
• Rabbit liver lysosomes adsorbed to Sepharose-anti-cathepsin M, but not to control Sepharose.

Conclusions:

• Membrane-bound cathepsin M is located on the outer surface of the lysosomal membrane.
• The monoclonal antibody provides a specific tool for identifying and localizing cathepsin M.