Dehydroalanine and Dehydrobutyrine in Aging and Cataractous Lenses

Protein post-translational modifications (PTMs) have been associated with aging and age-related diseases. PTMs are particularly impactful in long-lived proteins, such as those found in the ocular lens, because they accumulate with age. Two post-translational modifications that lead to protein-protein crosslinks in aged and cataractous lenses are dehydroalanine (DHA) and dehydrobutyrine (DHB); formed from cysteine/serine and threonine residues, respectively. The purpose of this study was to quantitate DHA and DHB in human lens proteins as a function of age and cataract status. Human lenses of various ages were divided into five donor groups: transparent lenses (18–22-year-old, 48–64-year-old, and 70–93-year-old) and cataractous human lenses of two age groups (48–64-year-old lenses, and 70–93-year-old lenses) and were subjected to proteomic analysis. Relative DHA and DHB peptide levels were quantified and compared to their non-modified peptide counterparts. For most lens proteins containing DHA or DHB, higher amounts of DHA- and DHB-modified peptides were detected in aged and cataractous lenses. DHA-containing peptides were classified into three groups based on abundance changes with age and cataract: those that (1) increased only in age-related nuclear cataract (ARNC), (2) increased in aged and cataractous lenses, and (3) decreased in aged lenses and ARNC. There was no indication that DHA or DHB levels were dependent on lens region. In most donor groups, proteins with DHA and DHB were more likely to be found among urea-insoluble proteins rather than among water- or urea-soluble proteins. DHA and DHB formation may induce structural effects that make proteins less soluble in water that leads to age-related protein insolubility and possibly aggregation and light scattering.

蛋白质翻译后修饰（PTMs）已被证实与衰老及衰老相关疾病存在关联。这类修饰对长寿蛋白的影响尤为显著，例如晶状体中的长寿蛋白，因为它们会随年龄增长不断积累。在衰老及白内障晶状体中，能够引发蛋白质间交联的两种翻译后修饰分别为脱氢丙氨酸（DHA）和脱氢丁酸（DHB），二者分别由半胱氨酸/丝氨酸残基与苏氨酸残基衍生而来。本研究旨在定量分析人晶状体蛋白中DHA与DHB的水平，并探究其与年龄及白内障状态的相关性。我们收集了不同年龄段的人晶状体，将其分为5个供体组：透明晶状体组（涵盖18~22岁、48~64岁及70~93岁三个亚组），以及两个年龄段的白内障人晶状体组（48~64岁组与70~93岁组），随后对所有样本开展蛋白质组学分析。我们对DHA与DHB修饰肽的相对水平进行了定量，并与对应未修饰肽的水平进行了比较。对于大多数携带DHA或DHB修饰的晶状体蛋白，在衰老及白内障晶状体中均可检测到更高水平的DHA和DHB修饰肽。根据修饰肽水平随年龄与白内障进展的丰度变化特征，含DHA的肽可被分为三类：（1）仅在年龄相关性核性白内障（ARNC）中水平升高；（2）在衰老及白内障晶状体中水平均升高；（3）在衰老晶状体及ARNC中水平降低。未发现DHA或DHB水平与晶状体区域存在相关性的证据。在大多数供体组中，携带DHA与DHB修饰的蛋白更易富集于尿素不溶性蛋白组分，而非水溶性或尿素可溶性蛋白组分。DHA与DHB的形成可能会诱导蛋白质结构改变，使其水溶性降低，进而引发衰老相关的蛋白质不溶性、聚集及光散射。