Characteristics of PPT1 and TPP1 enzymes in neuronal ceroid lipofuscinosis (NCL) 1 and 2 by dried blood spots (DBS) and leukocytes and their application to newborn screening.

Abstract

We first characterized PPT1 and TPP1 enzymes in dried blood spots (DBS), plasma/serum, and leukocytes/lymphocytes using neuronal ceroid lipofuscinosis (NCL) 1 and 2 patients and control subjects. PPT1 enzyme had only one acid form in control DBS, plasma/serum, and leukocytes/lymphocytes and showed deficient activities in these samples from NCL 1 patients. Conversely, TPP1 enzymes in control DBS and leukocytes/lymphocytes consisted of two forms, an acidic form and a neutral form, whereas serum TPP1 enzyme had only a neutral form. In control subjects, the optimal pH of PPT1 enzyme in DBS, plasma/serum, and leukocytes/lymphocytes was 4.5 to 5.0 in the acidic form, whereas TPP1 enzyme in control DBS and leukocytes/lymphocytes was pH 4.5 and 6.5, respectively. In NCL 1 and 2, both PPT1 and TPP1 enzyme activities in DBS, plasma, and leukocytes/lymphocytes were markedly reduced in acidic pH, whereas heterozygotes of NCL 1 and 2 in the acidic form showed intermediate activities between patients and control subjects. In neutral conditions, pH 6.0, the PPT1 enzyme activities in NCL 1 patients showed rather higher residual activities and intermediate activities in heterozygotes in NCL 1, which was probably caused by mutated proteins in three cases with NCL 1 patients. TPP1 enzyme activities at neutral pH 6.5 to 7.0 in DBS and leukocytes/lymphocytes showed higher enzyme activities in NCL 2 patients and heterozygotes. The reason for the increases of neutral TPP1 enzyme activities at pH 6.5 to 7.0 in NCL 2 DBS and leukocytes/lymphocytes, is obscure, but possibly caused by secondary activation of neutral TPP1 enzyme due to the absence of the acidic form. Interestingly, TPP1 activity in serum only consisted of a neutral form, no acidic form, and was not deficient in any NCL 2 patient. Therefore, we can diagnose NCL 1 patients by plasma/serum enzyme assay of PPT1, but not diagnose NCL 2 by serum TPP1 enzyme assay. A pilot study of newborn screening of NCL 1 and 2 has been established by more than 1000 newborn DBS assays. Using this assay system, we will be able to perform newborn screening of NCL 1 and 2 by DBS.

• Authors: Rina Itagaki, Masahiro Endo, Hiroko Yanagisawa, Mohammad Arif Hossain, Keiko Akiyama, Keiko Yaginuma, Takashi Miyajima, Chen Wu, Takeo Iwamoto, Junko Igarashi, Yu Kobayashi, Jun Tohyama, Kazuhiro Iwama, Naomichi Matsumoto, Haruo Shintaku, Yoshikatsu Eto
• Journal: Molecular genetics and metabolism (Mol Genet Metab) Vol. 124 Issue 1 Pg. 64-70 (05 2018) ISSN: 1096-7206 [Electronic] United States
• PMID: 29599076 (Publication Type: Journal Article, Research Support, Non-U.S. Gov't)
• Copyright: Copyright © 2018 Elsevier Inc. All rights reserved.

Chemical References

• Membrane Proteins
• Tripeptidyl-Peptidase 1
• Thiolester Hydrolases
• PPT1 protein, human
• Serine Proteases
• Aminopeptidases
• Dipeptidyl-Peptidases and Tripeptidyl-Peptidases
• TPP1 protein, human

Topics

• Adult
• Aminopeptidases (blood)
• Child
• Child, Preschool
• Dipeptidyl-Peptidases and Tripeptidyl-Peptidases (blood)
• Dried Blood Spot Testing (methods)
• Female
• Humans
• Hydrogen-Ion Concentration
• Infant, Newborn
• Leukocytes (chemistry)
• Male
• Membrane Proteins (blood)
• Mutation
• Neonatal Screening (methods)
• Neuronal Ceroid-Lipofuscinoses (diagnosis)
• Pilot Projects
• Serine Proteases (blood)
• Thiolester Hydrolases (blood)
• Tripeptidyl-Peptidase 1