Metal Complex Bioconjugates as Artificial Enzymes

这个无机、生物无机和有机金属化学项目的奖项支持爱荷华州立大学的Nenad M. Kostic博士合成具有前所未有的序列选择性和位点特异性的催化裂解蛋白质的试剂的研究。这些无机试剂补充酶在生物化学、分子生物学、药物化学和生物技术中的应用。本研究建立在以前nsf支持的工作的基础上，从配位立体化学和裂解反应的动力学和机制方面解释了Pd（II）和Pt（II）配合物作为裂解剂在序列选择性上的显著差异。目前的研究旨在开发一种新的Pd配合物裂解蛋白质的生物技术方法，并开发第二代含Pd的裂解试剂，以实现主链裂解的催化转换。具体目的是：(1)研究X- pro和X- pro - his片段中的残基X和局部蛋白质构象对中性pH下钯水络合物切割蛋白质的动力学和特异性的影响。然后将可剪切的片段插入融合蛋白中，以证明这些试剂可以在普通蛋白水解酶不足的情况下成功地切割融合蛋白；(2)开发两类新的钯-有机偶联试剂，其中钯水配合物的裂解多肽主链的能力与一个或两个附着的β -环糊精识别裂解位点附近芳侧链的能力相结合。这些同时识别-切割相互作用的立体化学要求将被研究并用于设计最佳试剂，用于切割X- X- pro -Y片段中的X- pro键，其中X或Y是苯丙氨酸，酪氨酸或色氨酸；(3)利用Pd-bis（β -环糊精）试剂，解决了产物抑制问题，实现了X- X- pro -Y段X- X- pro键在中性pH下的水解裂解催化翻转，其中X和Y都是芳香残基。通过与Invitrogen公司的合作，该公司将努力将这项研究的一些产品商业化，学生将学习工业科学，商业化和潜在的就业。这项研究将在主要期刊、研讨会和国际会议上广泛传播，并在化学家、生物化学家和分子生物学家的研究生专题课程中广泛传播。

This award in the Inorganic, Bioinorganic and Organometallic Chemistry program supports research by Dr. Nenad M. Kostic at Iowa State University to synthesize reagents that catalytically cleave proteins with unprecedented sequence-selectivity and site-specificity. These inorganic reagents complement enzymes in applications in biochemistry, molecular biology, pharmaceutical chemistry, and biotechnology. This research builds on previous NSF-supported work that explained a striking difference in the sequence-selectivity of Pd(II) and Pt(II) complexes as cleavage agents in terms of the coordination stereochemistry and the kinetics and mechanisms of the cleavage reactions. The current research aims to develop a new biotechnological method for protein cleavage by Pd complexes and also to develop a second generation of Pd-containing cleavage reagents that will achieve catalytic turnover in backbone cleavage. The specific objectives are to: (1) study the effects of the residue X in the X-Pro and X-Pro-His segments and of the local protein conformation on the kinetics and the specificity of protein cleavage at neutral pH by the palladium aqua complexes. The scissile segments will then be inserted into fusion proteins tp demonstrate that these reagents can successfully cleave fusion proteins, in cases in which common proteolytic enzymes are inadequate; (2) develop two classes of new palladium-organic conjugate reagents, in which the ability of the Pd aqua complexes to cleave the polypeptide backbone is combined with the ability of one or two attached beta-cyclodextrins to recognize aromatic side chains proximal to the cleavage site. The stereochemical requirements for these simultaneous recognition-cleavage interactions will be investigated and used in designing optimal reagents for cleaving the x-Pro bond in the X-x-Pro-y-Y segments in which X or Y or both are phenylalanine, tyrosine, or tryptophan; (3) solve the problem of product inhibition and achieve catalytic turnover in the hydrolytic cleavage at neutral pH of the x-Pro bond in the X-x-Pro-y-Y segments in which both X and Y are aromatic residues with the Pd-bis(beta-cyclodextrin) reagent. Through a partnership with Invitrogen Co., which will endeavor to commercialize some of the products of this research, students will learn about industrial science, commercialization, and potential employment. This research will be widely disseminated in leading journals, seminars and international meetings, and in a new graduate special-topics course for chemists, biochemists and molecular biologists.