Thermal denaturation of T4 gene 32 protein: effects of zinc removal and substitution.
Thermal denaturation of T4 gene 32 protein: effects of zinc removal and substitution.
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T4 基因 32 蛋白的热变性：锌去除和替代的影响。
DOI：
10.1021/bi00414a044
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发表时间：
1988
期刊：
影响因子：
2.9
通讯作者：
Sturtevant,JM
中科院分区：
文献类型：
--
作者：
Keating,KM;Ghosaini,LR;Giedroc,DP;Williams,KR;Coleman,JE;Sturtevant,JM
基因32蛋白（g32 P）是噬菌体T4的单链DNA结合蛋白，是一种锌金属蛋白。内在锌是蛋白质与ssDNA晶格协同结合所需的因素之一。我们已经使用差示扫描量热法来确定如何的热力学参数表征g32 P的变性的影响去除或取代的内在锌。在宽的浓度范围（1-10 mg/mL）内，天然Zn（II）蛋白在55 ℃的Tm下展开，相关的平均焓变为139 kcal mol-1。在相同条件下，无金属的脱辅基蛋白在以49 ℃为中心的相对较宽的温度范围内变性，平均焓变为84 kcal mol-1。取代锌（II）在g32 P由镉（II）或钴（II）不显着改变变性的焓，但会影响蛋白质的热稳定性。所有金属形式的g32 P结合到聚（dT）进行高度合作的变性转变，其特征在于不对称的差示扫描量热法峰与4-5 C相比，在tm的unliganded金属蛋白。从g32 P去除金属离子显著降低了与聚（dT）结合的协同性[Giedroc，D. P.，基廷，K。M.，威廉姆斯，K. R.，&科尔曼，JE（1987）Biochemistry 26，5251-5259]，并且推测作为其结果，apo-g32 P在结合到聚（dT）上时显示热转变的形状或中点没有变化。因此，当蛋白质与ssDNA协同复合时，与每个g32 P单体四面体配位的单个金属离子提供了最大的生物稳定性增加。在符合量热数据的雅阁中，我们认为，构象的变化或构象通量的减少必然是锌与g32 P结合的直接结果，而锌与g32 P的结合又使蛋白质与ssDNA.Cjfene 32的高度合作结合能够发生。这是DNA复制、修复和重组所需的化学计量量[综述见Chase和威廉姆斯（1986）]。g32 P与单链核酸结合的总自由能的大约三分之一来自与核酸晶格结合的相邻g32 P分子之间的协同蛋白质-蛋白质相互作用（Kelly et al.，1976年）。这些协同g32 P-g32 P相互作用部分依赖于一个完整的NH 2-末端“B”区（定义为残基1-21）。蛋白水解去除B区将结合ssDNA的协同性参数从约103降低到约104。
Revised Manuscript Received March 17, 1988 abstract: Gene 32 protein (g32P), the single-stranded (ss) DNA bindingprotein from bacteriophage T4, is a zinc metalloprotein. The intrinsic zinc is one of the factors required for the protein to bind cooperatively to a ssDNA lattice. We have used differential scanning calorimetry to determine how the thermodynamic parameters characterizing the denaturation of g32P are affected by removal or substitution of the intrinsic zinc. Over a wide concentration range (1-10 mg/mL), the native Zn (II) protein unfolds at a tm of 55 C with an associated mean enthalpy change of 139 kcal mol'1. Under the same conditions, the metal-free apoprotein denatures over a relatively broader temperaturerange centered at 49 C, with a mean enthalpy change of 84 kcal mol'1. Substitution of Zn (II) in g32P by either Cd (II) or Co (II) does not significantly change the enthalpy of denaturation but does affect the thermal stability of the protein. All metallo forms of g32P when bound to poly (dT) undergo highly cooperative denaturational transitions characterized by asymmetric differential scanning calorimetry peaks with increases in tm of 4-5 C compared to the unliganded metalloprotein. Removal of the metal ion from g32P significantly reduces the cooperativity of binding to poly (dT)[Giedroc, D. P., Keating, K. M., Williams, K. R., & Coleman, JE (1987) Biochemistry 26, 5251-5259], and presumably as a consequence of this, apo-g32P shows no change in either the shape or the midpoint of the thermal transition on binding to poly (dT). Thus, a single metal ion tetrahedrally coordinated to each g32P monomer provides the largest increase in the thermodynamicstability when the protein is cooperatively complexed with ssDNA. In accord with calorimetric data, we suggest that a change in the conformation or a reduction in conformational flux must occur as a direct consequence of the association of zinc with g32P which in turn enables highly cooperative binding of the protein to ssDNA.Cjfene 32 of bacteriophage T4 codes for a nucleic acid binding protein (g32P)! that is required in stoichiometric amounts for DNA replication, repair, and recombination [for a review, see Chase and Williams (1986)]. Approximately one-third of the overall free energy of g32P binding to sin-gle-stranded nucleic acids derives from cooperative protein-protein interactions between adjacent g32P molecules bound to the nucleic acid lattice (Kelly et al., 1976). These coop-erative g32P-g32P interactions are dependent in part upon an intact NH2-terminal “B” region (defined here as residues 1-21). Proteolytic removal of the B region reduces the co-operativity parameters for binding ssDNA from about 103 to
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T4 噬菌体基因 32：DNA 复制和重组中的结构蛋白
影响因子：
64.8
作者：
B. Alberts;L. Frey
通讯作者：
L. Frey
T4 基因 32 蛋白中锌的功能。
影响因子：
2.9
作者：
Giedroc,DP;Keating,KM;Williams,KR;Coleman,JE
通讯作者：
Coleman,JE
脱辅基、脱辅基磷酰基和金属碱性磷酸酶的差示扫描量热法。
DOI：
10.1016/s0021-9258(19)66932-2
发表时间：
1977
期刊：
The Journal of biological chemistry
影响因子：
--
作者：
J. Chlebowski;S. Mabrey
通讯作者：
S. Mabrey
影响因子：
5.6
作者：
CARROLL, RB;NEET, K;GOLDTHWAIT, DA
通讯作者：
GOLDTHWAIT, DA
影响因子：
5.6
作者：
B. Levine;D. Coffman;J. Thornton
通讯作者：
J. Thornton