Tissue amyloid P component in normal human dermis is non-covalently associated with elastic fiber microfibrils.
Tissue amyloid P component (TAP), a protein that crossreacts immunohistochemically with the normal plasma glycoprotein serum amyloid P component (SAP), is invariably associated with elastic fiber microfibrils in adult humans. We have investigated the nature of this association. Aliquots of minced, homogenized dermis, obtained following ethylenediamine tetraacetic acid (EDTA) separation of whole adult human skin, were extracted with different reagents, and the presence or absence of TAP in the pellet and in the supernatant following centrifugation was determined by SDS-PAGE and immunoblotting using anti-SAP antibodies. TAP was extractable from dermis using reagents which disrupt non-covalent bonds, including sodium dodecyl sulfate (SDS) and guanidine hydrochloride. TAP was not extracted by high molarity salt solutions, non-ionic detergents, or the reducing agents dithiothreitol and 2-mercaptoethanol. EDTA solution was similarly unsuccessful at eluting TAP from the dermal preparation, indicating that the association of TAP with elastic fiber microfibrils is not simply the result of Ca++-dependent binding. Collagenase solubilized some TAP, but this does not prove covalent linkage to elastic tissue of part of the TAP, because the apparent Mr of TAP extracted was identical to that of normal SAP subunits. We cannot completely exclude the possibility that a few subunits in each multimeric TAP molecule are covalently attached to the microfibrils. However, our findings that denaturing agents alone extracted most of the TAP from normal human dermis strongly suggest that the great majority of the dermal TAP is non-covalently bound to elastic fiber microfibrils. Thus TAP is not an integral constitutent of elastic fiber microfibrils.
Breathnach
S M
SM
Department of Medicine (Dermatology), Charing Cross and Westminster Medical School, London, U.K.
Pepys
M B
MB
Hintner
H
H
D000328:Adult; D004547:Elastic Tissue / Q000648:ultrastructure*; D006801:Humans; D015151:Immunoblotting; D007202:Indicators and Reagents; D000683:Serum Amyloid P-Component / Q000032:analysis*; D012867:Skin / Q000032:analysis* / Q000648:ultrastructure
D007202:Indicators and Reagents; D000683:Serum Amyloid P-Component