CHARACTERIZATION OF THE 33-KILODALTON MAJOR ALLERGEN OF PENICILLIUM-CITRINUM BY USING MOABS AND N-TERMINAL AMINO-ACID SEQUENCING

Background The 33 kD component has been identified as a major allergen of Penicillium citrinum, the most prevalent Penicillium species in th e Taipei area of Taiwan. Objective This study analyses the isoforms, a ntigenic cross-reactivity and the N-terminal amino acid sequence of th e 33 kD allergen of P. citrinum. Methods The composition of isoforms a nd antigenic cross-reactivity was analysed by SDS-PAGE and 2D-immunobl otting using MoAbs generated. The N-terminal sequence was analysed by using an automatic gas/liquid phase sequencer. Results Two MoAbs (55A and 34H) against the 33 kD allergen were generated in the present stud y. In addition to the 33 kD component, MoAb 34H also showed immunoblot reactivity to other components in the elude extract of P. citrinum. A nalysed by 2D-immunoblotting, at least six different isoforms of the 3 3 kD component with pi values ranging from 6.75 to greater than 7.0 we re shown to be reactive to both MoAbs and IgE antibodies in serum of a n asthmatic patient. Different immunoblot patterns were observed when both MoAbs were reacted with four different strains of P. citrinum use d in the present study. Among another six different Penicillium and fo ur different Aspergillus species tested, only an immunoblot reactivity of MoAb 55A to the 33 kD component of P. brevicompactum was observed. In 2D-immunoblotting, components of P. brevicompactum with an MW of a bout 33 kD and pI values similar to those of the 33 kD component of P. citrinum reacted with MoAb 55A and IgE antibodies in serum of the ast hmatic patient. The N-terminal amino acid sequence of the 33 kD compon ent of P. citrinum was determined to be ANVVQSNVP which was identical to the first 9 N-terminal amino acids of a heat-labile alkaline serine proteinase from P. citrinum. Conclusion Results obtained in the prese nt study suggest that the 33 kD major allergen of P. citrinum may be a n alkaline serine proteinase.