Identification of Hydroxyproline-Containing Proteins and Hydroxylation of Proline Residues in Rice

Abstract

The hydroxyproline-containing proteins (HCPs) among secretory and vacuolar proteins play important roles in growth and development of higher plants. Many hydroxyprolinerich glycoproteins (HRGPs), including Arabinogalactan proteins (AGPs), extensins (EXTs), and proline-rich proteins (PRPs), are identified as HCPs by bioinformatics approaches. The experimental evidence for validation of novel proline hydroxylation sites is vital for understanding their functional roles. In this study, the 62 HCPs containing 114 hydroxyproline (O, Hyp) residues were identified, and it was found that hydroxylation of proline residues in the HCPs could either constitute attachment sites for glycans or have other biological function in rice. The glycomodules of AO, OA, OG, VO, LO, and OE were abundant in the 62 HCPs. Further analysis showed that the 22 of 62 HCPs contained both signal peptides and transmembrane domains, and the 19 HCPs only contained transmembrane domains, while 21 HCPs contained neither. This study indicated the feasibility of mass spectrometry-based proteomics combined with bioinformatics approaches for the large-scale characterization of Hyp sites from complex protein digest mixtures. Furthermore, the expression of AGPs in rice was detected by using b-GlcY reagent and JIM13 antibody. The results displayed that the AGPs were widely distributed in different tissues and organs of rice, especially expressed highly in lateral root, pollen and embryo. In conclusion, our study revealed that the HCPs and Hyp residues in rice were ubiquitous and that these Hyps could be candidates for linking to glycans, which laid the foundation for further studying the functions of HCPs and hydroxylation of proline residues in rice.

Differential prolyl hydroxylation by six Physcomitrella prolyl-4 hydroxylases

“…In agreement with MS measurements of 114 Hyps from 62 glycoproteins in rice (Liang et al, 2020), we found alanine most often to precede Hyps. Valine was the second most frequent AA before Hyp in rice.…”

Glycosylated Polyhydroxyproline Is a Potent Antifreeze Molecule

“…Prior structural work on glycosides of Hyp focused mainly on Hyp-rich glycoproteins that are the major proteinaceous components of higher plant walls, such as Gp1 and extensin. , Predominant glycan structures are β-galactosides and α-arabinosides with varied chain lengths. CD spectra of these proteins indicated PPII structures; however, the specific effects of glycosylation were unclear since these natural peptides and proteins are typically isolable in only minuscule amounts and/or are heterogeneous in nature .…”

Estudio de defensinas de flores de cardos con potencial aplicación biotecnológica

“…Una de las modificaciones postraduccionales reversibles más importantes la constituye la hidroxilación de proteínas, siendo la prolina el principal residuo que sufre dicha modificación (Hu et al, 2010). Las proteínas que contienen hidroxiprolina juegan un papel clave en el crecimiento y desarrollo de las plantas superiores ya que serían sitios de oglicosilación correspondientes a modificaciones postraduccionales reversibles relevantes en diversos procesos fisiológicos (Liang et al, 2020). De hecho, se encontró que la hidroxiprolina es el aminoácido mayoritario en hidrolizados de pared celular (Deepak et al, 2010).…”