A 34-amino-acid peptide has been chemically synthesized based on a sequence from human alpha-fetoprotein. The purified peptide is active in anti-growth assays when freshly prepared in pH 7.4 buffer at 0.20 g/l, but this peptide slowly becomes inactive. This functional change is proven by mass spectrometry to be triggered by the formation of an intrapeptide disulfide bond between the two cysteine residues on the peptide. Interpeptide cross-linking does not occur. The active and inactive forms of the peptide have almost identical secondary structures as shown by circular dichroism (CD). Zinc ions bind to the active peptide and completely prevents formation of the inactive form. Cobalt(II) ions also bind to the peptide, and the UV-Vis absorption spectrum of the cobalt-peptide complex shows that: (1) a near-UV sulfur-to-metal-ion charge-transfer band had a molar extinction coefficient consistent with two thiolate bonds to Co(II); (2) the lowest-energy visible d-d transition maximum at 659 nm, also, demonstrated that the two cysteine residues are ligands for the metal ion; (3) the d-d molar extinction coefficient showed that the metal ion-ligand complex was in a distorted tetrahedral symmetry. The peptide has two cysteines, and it is speculated that the other two metal ion ligands might be the two histidines. The Zn(II)- and Co(II)-peptide complexes had similar peptide conformations as indicated by their ultraviolet CD spectra, which differed very slightly from that of the free peptide. Surprisingly, the cobalt ions acted in the reverse of the zinc ions in that, instead of stabilizing anti-growth form of the peptide, they catalyzed its loss. Metal ion control of peptide function is a saliently interesting concept. Calcium ions, in the conditions studied, apparently do not bind to the peptide. Trifluoroethanol and temperature (60 degrees C) affected the secondary structure of the peptide, and the peptide was found capable of assuming various conformations in solution. This conformational flexibility may possibly be related to the biological activity of the peptide.

The products produced by X irradiation of an oxygenated aqueous solution containing d(CpApTpG) were analyzed by NMR spectroscopy and mass spectrometry. Thirteen different base modifications were detected, including a novel product formed by the addition of oxygen to guanine. Seven different strand break products were identified, including strands having 5'-phosphoryl groups, 3'-phosphoryl groups and groups having 3'-phosphoglycolates as termini. The products produced in largest yield contained base modifications: Pyrimidine bases degraded to a formamido moiety, the 8-oxo-7,8-dihydroguanine (8-oxoguanine) lesion, and double base lesions in which both the 8-oxo-7,8-dihydroguanine lesion and a formamido remnant are present.

The Saccharomyces cerevisiae al homeodomain is expressed as a soluble protein in Escherichia coli when cultured in minimal medium. Nuclear magnetic resonance (NMR) spectra of previously prepared al homeodomain samples contained a subset of doubled and broadened resonances. Mass spectroscopic and NMR analysis demonstrates that the heterogeneity is largely due to a lysine misincorporation at the arginine (Arg) 115 site. Arg 115 is coded by the 5′‐AGA‐3′ sequence, which is quite rare in E. coli genes. Lower level mistranslation at three other rare arginine codons also occurs. The percentage of lysine for arginine misincorporation in al homeodomain production is dependent on media composition. The dnaY gene, which encodes the rare 5′‐AGA‐3′ tRNA ARG , was co‐expressed in E. coli with the al‐encoding plasmid to produce a homogeneous recombinant al homeodomain. Co‐expression of the dnaY gene completely blocks mistranslation of arginine to lysine during al overexpression in minimal media, and homogeneous protein is produced.

A new glycoamidase, peptide-N 4-(N-acetyl-β-d-glucosaminyl)asparagine amidase (PNGase) At, was discovered in the eukaryote Aspergillus tubigensis. The enzyme was purified to homogeneity, and the DNA sequence was determined by cloning in Escherichia coli. Over 80% of the deduced amino acid sequence was verified independently by Edman analysis and/or electrospray ionization-mass spectrometry of protease fragments of native PNGase At. This glycoamidase contains 12 potential asparagine-linked glycosylation sites, of which at least 9 sites are occupied with typical high mannose oligosaccharides. PNGase At consists of two non-identical glycosylated subunits that are derived from a single polypeptide gene precursor. Evidence is presented suggesting that autocatalysis is involved in subunit formation. PNGase At is an important new tool for analysis of asparagine-linked glycans; it can hydrolyze a broad range of glycopeptides, including those with core-linked α1→6 or α1→3 fucose, under conditions not favorable with existing glycoamidases.

C-Phycocyanin has been purified from Synechococcus lividus (SyI), which grows between 66 and 73°C at the highest temperature of any cyanobacterium and of any biliprotein-containing organism. The protein was examined at its physiological temperature, 70°C, in order to compare with its properties at 20°C. The protein was found to have virtually identical properties of light harvesting, bilin conformation, fluorescence, and secondary protein structure at both temperatures. The unique absorption maximum of SyI protein was maintained at 70°C. Using the bilins as built-in reporter groups, the response is to resist, over a wide temperature range (10–80°C), the denaturation that would occur in mesophiles, rather than exhibit special adaptation to 70°C. C-Phycocyanin from another thermophile, S. lividus (SyIII), behaved differently. The SyI protein was `temperature-resistant', and the other `cold-dissociated'. In addition, an early assembly step toward the phycobilisome is the formation of monomers (αβ) from bilin-bearing, α and β polypeptides. Unlike larger aggregates, C-phycocyanin monomers from SyI are found to denature between 60°C and 70°C. The instability of monomers at their physiological temperature suggests that they either rapidly aggregate to avoid denaturation, or they are protected from denaturation by some agent. Monomers of the thermophile were, however, much more stable against high temperature than the monomers from mesophiles. The structure of SyI C-phycocyanin and its phycobilisomes were shown to be similar to those of mesophiles.