The most stabilizing point mutation was D27F, which is located in the active site of the protein, rendering it inactive.

PopMusic shows also strong performance with highly statistically significant 1’ = 0.55 between theory and experiment, however the limitation of this method is that it can consider only single point mutations .

Computational identification of stabilizing single point mutations

All possible single point mutations of DHFR (159 * 19 = 3,021) were simulated with the Monte Carlo protein unfolding simulation protocol.

The distribution of predicted melting temperatures (averaged over the 3 metrics) for all 3021 point mutants is shown in Fig.

However using MCPU we were able to efficiently explore stabilities of all possible point mutants for an essential enzyme of a typical size (159 amino acids) in a manageable amount of computational time (approx.

We cloned, expressed, and purified the 23 single point mutants of DHFR listed in 81 Table, as well as the multiple mutants listed in Table 1 (see Materials and Methods).

Of the selected 22 single point mutations , 10 mutations were stabilizing, according to their Tm or Cm values (82 Table).

Here, we use a Monte Carlo protein unfolding approach (MCPU) with an all-atom simulation method and knowledge-based potential developed earlier in our lab [16,30,31] to simulate unfolding and predict melting temperatures for all possible single point mutants of E. coli Dihydrofolate Reductase (DHFR).

Single point mutations of DHFR were constructed based on a two-step PCR-mutagenesis strategy [70], in which the template for the PCR is the plasmid of WT DHFR.

In the current work, we only consider resistance to a single drug, or more precisely, to treatments with a drug or drug combination to which resistance can be generated by the accumulative acquisition of a set of point mutations .

Denote by i the genotype of a cancer cell if it has acquired 1' point mutations (i = 0, 1).

We assume that one point mutation is sufficient to confer high levels of resistance to the maximum possible concentration of drugs administered during therapy.

We denote the genotypes of cells by the number of acquired point mutations : the wild type ‘0’ and the resistant type ‘1’ (see Materials & Methods for a detailed description of the model).

Later on, we will extend this simple model to more realistic cases with multiple cell types and with multiple compartments, where multiple point mutations can successively accrue to confer resistance to exceedingly high drug concentrations [54].

Four cell lines [SH-SY5Y, LAN-6, SMS-KCN, and SK-N-BE(2)] were selected for further studies because of different point mutations in ALK, p53 status, RTK expression, morphology, and growth characteristics.

Four cell lines [SH-SY5Y, LAN-6, SMS-KCN, and SK-N-BE(2)] were selected for further studies because of their different point mutations in ALK, p53 status, RTK expression, morphology, and growth patterns.

Point mutations in the RTK, ALK, are the primary cause of familial neuroblastoma and account for 8—12% of sporadic neuroblastomas [15].

This observation, combined with evidence from our MD simulations that BsFtsZ pocket scores are on par with the scores of point mutations that confer PC190723 resistance in S. aureus (Fig.

Single point mutations were introduced into the S. aureus crystal structure with the mutate residue VMD modeling extension [37].

Furthermore, single point mutations conferring resistance to PC190723 in S. aureus were identified within ftsZ, suggesting that PC190723 specifically binds to S. aureus FtsZ (SaFtsZ).