Small leucine-rich repeat protein (SLRP) family members contain conserved leucine-rich repeat motifs flanked by highly variable N- and C-terminal regions. Most class II and III SLRPs have tyrosine-rich N-terminal regions and some of these are sulfated. However, the evolutionary origin and conservation of the tyrosine-rich and acidic terminal regions remain undetermined. In this study, we present the most comprehensive multiple sequence alignment (MSA) analyses of all eight class II and III SLRPs to date. Based on the level of conservation of tyrosine residues and adjacent sequences, we predict which tyrosine residues are most likely to be sulfated in the terminal regions of human class II and III SLRPs. Using this novel approach, we predict a total of 22 tyrosine sulfation sites in human SLRPs, of which only 8 sites had been experimentally identified in mammals. Our analyses suggest that sulfation-prone, tyrosine-rich and acidic terminal regions of the class II and III SLRPs emerged via convergent evolution at different stages of vertebrate evolution, coinciding with significant evolutionary events including the development of endochondral bones and articular cartilage, the aquatic to terrestrial transition, and the formation of an amnion. Our study suggests that selective pressures due to changes in life conditions led to the formation of sulfotyrosine-rich and acidic terminal regions. We believe the independent emergence and evolution of sulfotyrosine-rich and acidic N- and C-terminal regions have provided each class II and III SLRP member with novel vital functions required to develop new specialized extracellular matrices and tissues in vertebrate species.

中文翻译：

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II类和III类富含亮氨酸的小硫酸化重复序列的硫酸化倾向酪氨酸富集和酸性N-和C-末端延伸的起源和发展为脊椎动物的结缔组织进化提供了启示。

小的富含亮氨酸的重复蛋白（SLRP）家族成员包含保守的富含亮氨酸的重复基序，其侧翼是高度可变的N和C端区域。大多数II类和III类SLRP具有富酪氨酸的N末端区域，其中一些被硫酸化。但是，酪氨酸丰富和酸性终端区的进化起源和保存仍未确定。在这项研究中，我们提出了迄今为止所有八种II类和III类SLRP的最全面的多序列比对（MSA）分析。基于酪氨酸残基和相邻序列的保守水平，我们预测哪些酪氨酸残基最有可能在人类II类和III类SLRP的末端区域被硫酸化。使用这种新颖的方法，我们可以预测人类SLRP中共有22个酪氨酸硫酸化位点，在哺乳动物中，只有8个位点已通过实验鉴定。我们的分析表明，在脊椎动物进化的不同阶段，趋于硫酸化的，富含酪氨酸的和II类和III类SLRP的末端区域通过趋同进化而出现，同时也伴随着重大的进化事件，包括软骨内骨骼和关节软骨的发育，向地面过渡，并形成羊膜。我们的研究表明，由于生活条件的变化而产生的选择性压力导致形成富含硫代酪氨酸和酸性的末端区域。