Top-down HPLC-ESI-MS detection of S-glutathionylated and S-cysteinylated derivatives of cystatin B and its 1-53 and 54-98 fragments in whole saliva of human pre-term newborns

Iavarone F;CABRAS, TIZIANA;Pisano E;SANNA, MARIA TERESA;Nemolato S;Vento G;Tirone C;Romagnoli C;Cordaro M;FANOS, VASSILIOS;FAA, GAVINO;MESSANA, IRENE;Castagnola M.

2013-01-01

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Abstract

Analysis by a HPLC-ESI-MS top-down proteomic platform of specimens of human preterm newborn whole saliva evidenced high relative amounts of cystatin B and its S-glutathionylated,S-cysteinylated, and S-S 2-mer (on Cys(3)) derivatives, decreasing as a function of postconceptional age (PCA). The percentage of S-unmodified cystatin B was higher than the S-modified isoforms in the early PCA period, differently from adults where cystatin B was detectable only as S-modified derivatives. The percentage of S-modified derivatives increased as a function of PCA, reaching at the normal term of delivery values similar to those determined in at-term newborns, babies, and adults. Moreover, in the early PCA period, high relative amounts of the 1-53 and 54-98 cystatin B fragments were detected, decreasing as a function of PCA and disappearing at the normal term of delivery. In agreement with intact cystatin B, fragment 1-53 was detectable as S-unmodified and S-modified derivatives, and their percentages changed accordingly with the percentages of intact proteins, suggesting that the fragmentation process could be subsequent to and independent from the S-modification of the protein. This study highlights specific enzymatic activity in the oral cavity of preterm newborns not present in at-term newborns and adults, which can be a clue to specialized pathways occurring during fetal oral development.