Leukocyte Phosphomannomutase and Phosphomannose Isomerase Activity in an Indian Cohort
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Leukocyte Phosphomannomutase and Phosphomannose Isomerase Activity in an Indian Cohort Mihika B. Dave1 • Alpa J. Dherai1
•
Vrajesh P. Udani2 • Tester F. Ashavaid1
Received: 26 May 2020 / Accepted: 14 October 2020 Ó Association of Clinical Biochemists of India 2020
Abstract Advances in molecular sequencing technology has increased the diagnostic yield for Congenital disorder of glycosylation (CDG). However, novel variants or those of uncertain significance (vus) often pose a challenge and in such cases confirmed diagnosis can be warranted through enzyme analysis of these defects. We thus, aimed to optimize leukocyte-based enzyme assays for first two enzymes involved in N-glycosylation pathway i.e. Phosphomannomutase (PMM) and Phosphomannose isomerase (MPI). Study population comprised of 50 healthy non-alcoholic adults and 20 pediatric controls. Leukocyte enzyme activity was measured by monitoring the conversion of NADP to NADPH at 340 nm. The conditions were optimized and precision was assessed for both low and normal activity leukocyte controls. Enzyme activities for PMM and MPI in healthy individuals were measured in the range 1.6–3.9 and 7–20 nmol/min/mg protein respectively and did not vary with age and gender. The precision for both PMM and MPI showed %CV of 19.9 and 19.8 respectively. The enzyme activity in leukocyte pellet was found to be stable for up to 9 months when stored at -80 °C. The enzyme assays are optimized for PMM and MPI and can be used for evaluation of CDG patients in India.
& Alpa J. Dherai [email protected]; [email protected] 1
Department of Biochemistry, P. D. Hinduja Hospital & MRC, Veer Savarkar Marg, Mahim, Mumbai 400016, India
2
Department of Pediatric Neurology, P. D. Hinduja Hospital & MRC, Veer Savarkar Marg, Mahim, Mumbai 400016, India
Keywords Congenital disorder of glycosylation Leukocytes Phosphomannomutase Phosphomannose isomerase Variant of uncertain significance
Introduction N-glycosylation is a site-specific, multi-step enzymatic process that occurs in the cytoplasm, endoplasmic reticulum, and golgi apparatus [1]. The biosynthesis of all N-glycans occurs coherently and is essential in varied biological processes like protein folding, cell–cell interaction etc. Defects in glycosylation are implicated in several genetic and chronic diseases and complete absence of N-glycans is fatal [1]. Congenital disorders of glycosylation (CDG) are a group of over 100 different clinically and genetically heterogeneous glycosylation defects and * 50% of them are related to N-glycosylation disorders [2]. These glycosylation defects are distinct disorders and are not influenced by non-enzymatic glycation as seen in hyperglycemia. They are detected by transferrin isoform analysis wherein an increased disialo and/or asialo transferrin suggests type I defects while increase in tri, di, mono and/or asialo transferrin is indicative of type II N-glycosylation disorders [3]. Phosphomannomutase (PMM) and Phosphomannose isomerase (MPI) are primar
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