Structural basis for prolidase deficiency disease mechanisms

Abstract

All refined structure coordinates as well as the corresponding structure factor amplitudes have been deposited in the PDB under the accession numbers 5MBY, 5MBZ, 5MC0, 5MC1, 5MC2, 5MC3, 5MC4, 5MC5, 6H2P, 6H2Q.

Co‐expression with chaperones can affect protein 3D structure as exemplified by loss‐of‐function variants of human prolidase

“…Several studies showed the potential increase in total prolidase activity in cultured fibroblasts [41][42][43][44] and various therapeutic therapies were investigated [9,[45][46][47][48][49][50][51], but to date no efficient treatment was introduced. In recent years, we have structurally characterized wt prolidase [25] and a series of eight single amino acid substitution or deletions [10] and we could show that one of the major causes of loss of function (LOF) was structural destabilization induced by bulky side chains introduced in places of small ones. Previously, Besio et al showed that the induction of the chaperonins Hsp90 and Hsp70 in cultured fibroblasts derived from patients leads to partial rescue of prolidase activity.…”

“…A single peak corresponding to prolidase was collected, concentrated to~50 mgÁmL À1 , and used for crystallization trials or aliquoted and flash-frozen in liquid nitrogen (LN 2 ) for later use. Prolidase expression in Rosetta (DE3) cells followed the same procedure and was previously described [10,25].…”

“…Purified prolidase was crystallized as previously reported [10,25]. Protein solution in storage buffer (13-18 mgÁmL À1 ) was set up in sitting drop 96-well Intelli low-profile crystallization plates as a 1 : 1 mix with reservoir solution (10 mM sodium tetraborate and 720-1050 mM sodium citrate, pH: 7.5-8.5).…”

“…Relevant data collection and refinement statistics are summarized in Table S1 for the structures derived from Arctic Express cells. For the structures derived from Rosetta cells, the relevant parameters have been reported previously [10,25].…”

“…A series of eight mutations that result in single amino acid deletion or substitution has recently been analyzed by X-ray crystallography. In three of the resulting crystal structures, namely in the structures of the Ser202Phe, Gly287Asn, and Gly448Arg variants of the enzyme, significant level of protein disorder in the region winding around the active site was observed [10].…”